Use of pyrazolopyrimidine derivatives for the treatment of pi3k-delta related disorders

ABSTRACT

or pharmaceutically acceptable salts thereof.

This application is a continuation of U.S. Ser. No. 14/193,481, filedFeb. 28, 2014, which claims the benefit of priority of U.S. ProvisionalAppl. No. 61/771,480, filed Mar. 1, 2013, which is incorporated hereinby reference in its entirety.

TECHNICAL FIELD

The present application provides methods of treating PI3Kδ relateddisorders using pyrazolopyrimidine derivatives.

BACKGROUND

The phosphoinositide 3-kinases (PI3Ks) belong to a large family of lipidsignaling kinases that phosphorylate phosphoinositides at the D3position of the inositol ring (Cantley, Science, 2002,296(5573):1655-7). PI3Ks are divided into three classes (class I, II,and III) according to their structure, regulation and substratespecificity. Class I PI3Ks, which include PI3Kα, PI3Kβ, PI3Kγ, andPI3Kδ, are a family of dual specificity lipid and protein kinases thatcatalyze the phosphorylation of phosphatidylinosito-4,5-bisphosphate(PIP₂) giving rise to phosphatidylinosito-3,4,5-trisphosphate (PIP₃).PIP₃ functions as a second messenger that controls a number of cellularprocesses, including growth, survival, adhesion and migration. All fourclass I PI3K isoforms exist as heterodimers composed of a catalyticsubunit (p110) and a tightly associated regulatory subunit that controlstheir expression, activation, and subcellular localization. PI3Kα,PI3Kβ, and PI3Kδ associate with a regulatory subunit known as p85 andare activated by growth factors and cytokines through a tyrosinekinase-dependent mechanism (Jimenez, et al., J Biol Chem., 2002,277(44):41556-62) whereas PI3Kγ associates with two regulatory subunits(p101 and p84) and its activation is driven by the activation ofG-protein-coupled receptors (Brock, et al., J Cell Biol., 2003,160(1):89-99). PI3Kα and PI3Kβ are ubiquitously expressed. In contrast,PI3Kγ and PI3Kδ are predominantly expressed in leukocytes(Vanhaesebroeck, et al., Trends Biochem Sci., 2005, 30(4):194-204).

The differential tissue distribution of the PI3K isoforms factors intheir distinct biological functions. Genetic ablation of either PI3Kα orPI3Kβ results in embryonic lethality, indicating that PI3Kα and PI3Kβhave essential and non-redundant functions, at least during development(Vanhaesebroeck, et al., 2005). In contrast, mice which lack PI3Kγ andPI3Kδ are viable, fertile and have a normal life span although they showan altered immune system. PI3Kγ deficiency leads to impaired recruitmentof macrophages and neutrophils to sites of inflammation as well asimpaired T cell activation (Sasaki, et al., Science, 2000,287(5455):1040-6). PI3Kδ-mutant mice have specific defects in B cellsignaling that lead to impaired B cell development and reduced antibodyresponses after antigen stimulation (Clayton, et al., J Exp Med. 2002,196(6):753-63; Jou, et al., Mol Cell Biol. 2002, 22(24):8580-91;Okkenhaug, et al., Science, 2002, 297(5583):1031-4).

The phenotypes of the PI3Kγ and PI3Kδ-mutant mice suggest that theseenzymes may play a role in inflammation and other immune-based diseasesand this is borne out in preclinical models. PI3Kγ-mutant mice arelargely protected from disease in mouse models of rheumatoid arthritis(RA) and asthma (Camps, et al., Nat Med. 2005, 11(9):936-43; Thomas, etal., Eur J Immunol. 2005, 35(4):1283-91). In addition, treatment ofwild-type mice with a selective inhibitor of PI3Kγ was shown to reduceglomerulonephritis and prolong survival in the MRL-lpr model of systemiclupus nephritis (SLE) and to suppress joint inflammation and damage inmodels of RA (Barber, et al., Nat Med. 2005, 11(9):933-5; Camps, et al.,2005). Similarly, both PI3Kδ-mutant mice and wild-type mice treated witha selective inhibitor of PI3Kδ have been shown to have attenuatedallergic airway inflammation and hyper-responsiveness in a mouse modelof asthma (Ali, et al., Nature. 2004, 431(7011):1007-11; Lee, et al.,FASEB J. 2006, 20(3):455-65) and to have attenuated disease in a modelof RA (Randis, et al., Eur. J. Immunol., 2008, 38(5):1215-24).

B cell proliferation has shown to play a major role in the developmentof inflammatory autoimmune diseases (Puri, Frontiers in Immunology(2012), 3(256), 1-16; Walsh, Kidney International (2007) 72, 676-682).For example, B cells support T-cell autoreactivity, an importantcomponent of inflammatory autoimmune dieases. Once activated andmatured, B cells can traffic to sites of inflammation and recruitinflammatory cells or differentiate to plasmablasts. Thus, activity ofB-cells can be affected by targeting B-cell stimulatory cytokines,B-cell surface receptors, or via B-cell depletion. Rituximab—an IgGi κmouse/human chimeric monoclonal antibody directed against the B-cellsurface receptor CD20—has been shown to deplete CD20+ B cells. Use ofrituximab has been shown to have efficacy in treating idiopathicthrombocytopenic purpura, autoimmune hemolytic anemia, or vasculitis.For example, treatment with rituximab resulted in remission of thedisease in patients suffering from anti-neutrophil cytoplasm antibodyassociated (ANCA) systemic vasculitis (AASV) with demonstratedperipheral B-cell depletion (Walsh, 2007; Lovric, Nephrol DialTransplant (2009) 24: 179-185). Similarly, a complete response wasreported in one-third to two-thirds of patients having mixedcryoglobulinemia vasculitis after treatment with rituximab, includingpatients who presented with a severe form of vasculitis that wasresistant or intolerant to other treatments (Cacoub, Ann Rheum Dis 2008;67:283-287). Similarly, rituximab has been shown to have efficacy intreating patients with idiopathic thrombocytopenic purpura (or immunethrombocytopenic purpura) (Garvey, British Journal of Haematology,(2008) 141, 149-169; Godeau, Blood (2008), 112(4), 999-1004; Medeo,European Journal of Haematology, (2008) 81, 165-169) and autoimmunehemolytic anemia (Garvey, British Journal of Haematology, (2008) 141,149-169).

PI3Kδ signaling has been tied to B cell survival, migration, andactivation (Puri, Frontiers in Immunology, 2012, 3(256), 1-16, at pages1-5; and Clayton, J Exp Med, 2002, 196(6):753-63). For example, PI3Kδ isrequired for antigen-dependent B-cell activation driven by B cellreceptor. By blocking B-cell adhesion, survival, activation, andproliferation, PI3Kδ inhibition can impair the ability of B cells toactivate T cells, preventing their activation and reducing secreation ofautoantibodies and pro-inflammatory cytokines. Hence, by their abilityto inhibit B cell activation, PI3Kδ inhibitors would be expected totreat B cell mediated diseases that were treatable by similar methodssuch as B cell depletion by rituximab. Indeed, PI3Kδ inhibitors havebeen shown to be useful mouse models of various autoimmune diseases thatare also treatable by rituximab such as arthritis (Puri (2012)).Further, innate-like B cells, which are linked to autoimmunity aresensitive to PI3Kδ activity, as MZ and B-1 cells are nearly absent inmice lacking the p1106 gene (Puri (2012). PI3Kδ inhibitors can reducetrafficking of and activation of MZ and B-1 cells, which are implicatedin autoimmune diseases.

In addition to their potential role in inflammatory diseases, all fourclass I PI3K isoforms may play a role in cancer. The gene encoding p110αis mutated frequently in common cancers, including breast, prostate,colon and endometrial (Samuels, et al., Science, 2004, 304(5670):554;Samuels, et al., Curr Opin Oncol. 2006, 18(1):77-82). Eighty percent ofthese mutations are represented by one of three amino acid substitutionsin the helical or kinase domains of the enzyme and lead to a significantupregulation of kinase activity resulting in oncogenic transformation incell culture and in animal models (Kang, et al., Proc Natl Acad Sci USA.2005, 102(3):802-7; Bader, et al., Proc Natl Acad Sci USA. 2006,103(5):1475-9). No such mutations have been identified in the other PI3Kisoforms although there is evidence that they can contribute to thedevelopment and progression of malignancies. Consistent overexpressionof PI3Kδ is observed in acute myeloblastic leukemia (Sujobert, et al.,Blood, 2005, 106(3):1063-6) and inhibitors of PI3Kδ can prevent thegrowth of leukemic cells (Billottet, et al., Oncogene. 2006,25(50):6648-59). Elevated expression of PI3Kγ is seen in chronic myeloidleukemia (Hickey, et al., J Biol Chem. 2006, 281(5):2441-50).Alterations in expression of PI3Kβ, PI3Kγ and PI3Kδ have also beenobserved in cancers of the brain, colon and bladder (Benistant, et al.,Oncogene, 2000, 19(44):5083-90; Mizoguchi, et al., Brain Pathol. 2004,14(4):372-7; Knobbe, et al., Neuropathol Appl Neurobiol. 2005,31(5):486-90). Further, these isoforms have all been shown to beoncogenic in cell culture (Kang, et al., 2006).

For these reasons, there is a need to develop new PI3K inhibitors thatcan be used inflammatory disorders, autoimmune diseases and cancer. Thisinvention is directed to this need and others.

SUMMARY

The present invention provides methods of idiopathic thrombocytopenicpurpura, autoimmune hemolytic anemia, vasculitis, systemic lupuserythematosus, lupus nephritis, pemphigus, membranous nephropathy,chronic lymphocytic leukemia (CLL), Non-Hodgkin lymphoma, hairy cellleukemia, Mantle cell lymphoma, small lymphocytic lymphoma, follicularlymphoma, lymphoplasmacytic lymphoma, extranodal marginal zone lymphoma,Hodgkin's lymphoma, Waldenstrom's macroglobulinemia, prolymphocyticleukemia, acute lymphoblastic leukemia, myelofibrosis, mucosa-associatedlymphatic tissue (MALT) lymphoma, mediastinal (thymic) large B-celllymphoma, lymphomatoid granulomatosis, splenic marginal zone lymphoma,primary effusion lymphoma, intravascular large B-cell lymphoma, plasmacell leukemia, extramedullary plasmacytoma, smouldering myeloma (akaasymptomatic myeloma), monoclonal gammopathy of undeterminedsignificance (MGUS), activated B-cell like (ABC) diffuse large B celllymphoma, or germinal center B cell (GCB) diffuse large B cell lymphomain a patient, comprising administering to said patient a therapeuticallyeffective amount of a compound of Formula I:

or a pharmaceutically acceptable salt thereof, wherein R², R³, R⁴, R⁵,and Cy are defined infra.

The present invention also provides a compound described herein, or apharmaceutically acceptable salt thereof, for use in any of the methodsdescribed herein.

The present invention further provides use of a compound describedherein, or a pharmaceutically acceptable salt thereof, for themanufacture of a medicament for use in any of the methods describedherein.

DESCRIPTION OF DRAWINGS

FIG. 1 depicts the crystal structure of the compound of Example 269.

FIG. 2 depicts the tumor inhibiting effect of twice daily doses ofExample 347 at 0.3, 1, 3, or 10 mg/kg for 14 days in a Pfeiffer humantumor xenograft model of diffuse large B-cell lymphoma (y-axis is tumorvolume (mm³±SEM); x-axis is days post implantation).

DETAILED DESCRIPTION

The present invention provides a method of treating idiopathicthrombocytopenic purpura, autoimmune hemolytic anemia, vasculitis,systemic lupus erythematosus, lupus nephritis, pemphigus, membranousnephropathy, chronic lymphocytic leukemia (CLL), Non-Hodgkin lymphoma,hairy cell leukemia, Mantle cell lymphoma, small lymphocytic lymphoma,follicular lymphoma, lymphoplasmacytic lymphoma, extranodal marginalzone lymphoma, Hodgkin's lymphoma, Waldenstrom's macroglobulinemia,prolymphocytic leukemia, acute lymphoblastic leukemia, myelofibrosis,mucosa-associated lymphatic tissue (MALT) lymphoma, mediastinal (thymic)large B-cell lymphoma, lymphomatoid granulomatosis, splenic marginalzone lymphoma, primary effusion lymphoma, intravascular large B-celllymphoma, plasma cell leukemia, extramedullary plasmacytoma, smoulderingmyeloma (aka asymptomatic myeloma), monoclonal gammopathy ofundetermined significance (MGUS), activated B-cell like (ABC) diffuselarge B cell lymphoma (ABC-DLBCL, or germinal center B cell (GCB)diffuse large B cell lymphoma (GCB-DLBCL) in a patient, comprisingadministering to said patient a therapeutically effective amount of acompound of Formula I:

or a pharmaceutically acceptable salt thereof, wherein:

R² is C₁₋₆ alkyl or C₁₋₆ haloalkyl;

R⁴ is halo, OH, CN, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxy, or C₁₋₄haloalkoxy;

R⁵ is halo, OH, CN, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄ alkoxy, or C₁₋₄haloalkoxy; Cy is selected from C₃₋₇ cycloalkyl, 4-7 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl, each of which isoptionally substituted with 1, 2, 3, or 4 independently selected R³groups;

each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo,CN, NO₂, C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₁₋₆ haloalkyl,OR^(a1), SR^(a1), C(═O)R^(b1), C(═O)NR^(c1)R^(d1), C(═O)OR^(a1),OC(═O)R^(b1), OC(═O)NR^(c1)R^(d1), NR^(c1)R^(d1), NR^(c1)C(═O)R^(b1),NR^(c1)C(═O)OR^(b1), NR^(c1)C(═O)NR^(c1)R^(d1), C(═NR^(e))R^(b1),C(═NR^(e))NR^(c1)R^(d1), NR^(c1)C(═NR^(e))NR^(c1)R^(d1),NR^(c1)S(═O)R^(b1), NR^(c1)S(═O)₂NR^(c1)R^(d1), S(═O)R^(b1),S(═O)₂R^(b1), and S(═O)₂NR^(c1)R^(d1); wherein said C₁₋₆ alkyl, C₂₋₆alkenyl, C₂₋₆ alkynyl are each optionally substituted with 1, 2, 3, or 4independently selected R¹¹ groups;

each Cy¹ is independently selected from C₃₋₇ cycloalkyl, 4-7 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl, each of which isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups;

each R^(a1), R^(c1), and R^(d1) is independently selected from H, C₁₋₆alkyl, C₁₋₆ haloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇ cycloalkyl, 4-7membered heterocycloalkyl, phenyl, and 5-6 membered heteroaryl; whereinsaid C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇ cycloalkyl, 4-7membered heterocycloalkyl, phenyl and 5-6 membered heteroaryl are eachoptionally substituted with 1, 2, or 3 independently selected R¹¹groups;

each R^(b1) is independently selected from C₁₋₆ alkyl, C₁₋₆ haloalkyl,C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇ cycloalkyl, 4-7 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl; wherein said C₁₋₆alkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇ cycloalkyl, 4-7 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl are eachoptionally substituted with 1, 2, or 3 independently selected R¹¹groups;

or R^(c1) and R^(d1) together with the N atom to which they are attachedform a 4-, 5-, 6-, or 7 membered heterocycloalkyl group, which isoptionally substituted with —OH or C₁₋₃ alkyl;

each R^(e) is independently selected from H, CN, OH, C₁₋₄ alkyl, andC₁₋₄ alkoxy; and

each R¹¹ is independently selected from OH, NO₂, CN, halo, C₁₋₃ alkyl,C₂₋₃ alkenyl, C₂₋₃ alkynyl, C₁₋₃ haloalkyl, cyano-C₁₋₃ alkyl, HO—C₁₋₃alkyl, C₁₋₃ alkoxy-C₁₋₃ alkyl, C₃₋₇ cycloalkyl, C₁₋₃ alkoxy, C₁₋₃haloalkoxy, amino, C₁₋₃ alkylamino, di(C₁₋₃ alkyl)amino, thio, C₁₋₃alkylthio, C₁₋₃ alkylsulfinyl, C₁₋₃ alkylsulfonyl, carbamyl, C₁₋₃alkylcarbamyl, di(C₁₋₃ alkyl)carbamyl, carboxy, C₁₋₃ alkylcarbonyl, C₁₋₄alkoxycarbonyl, C₁₋₃ alkylcarbonylamino, C₁₋₃ alkylsulfonylamino,aminosulfonyl, C₁₋₃ alkylaminosulfonyl, di(C₁₋₃ alkyl)aminosulfonyl,aminosulfonylamino, C₁₋₃ alkylaminosulfonylamino, di(C₁₋₃alkyl)aminosulfonylamino, aminocarbonylamino, C₁₋₃alkylaminocarbonylamino, and di(C₁₋₃ alkyl)aminocarbonylamino.

In some embodiments, R² is C₁₋₃ alkyl or C₁₋₃ fluoroalkyl. In someembodiments, R² is methyl, ethyl, or 2,2-difluoromethyl. In someembodiments, R² is methyl. In some embodiments, R² is ethyl.

In some embodiments, R⁴ is halo, CN, or C₁₋₃ alkyl. In some embodiments,R⁴ is F, Cl, CN, or methyl. In some embodiments, R⁴ is F. In someembodiments, R⁴ is Cl. In some embodiments, R⁴ is CN. In someembodiments, R⁴ is methyl.

In some embodiments, R⁵ is halo, CN, or C₁₋₃ alkyl. In some embodiments,R⁵ is Cl, CN, or methyl. In some embodiments, R⁵ is Cl. In someembodiments, R⁵ is CN. In some embodiments, R⁵ is methyl.

In some embodiments, Cy is selected from C₃₋₆ cycloalkyl, 4-6 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl, each of which isoptionally substituted with 1, 2, 3, or 4 independently selected R³groups. In some embodiments, Cy is 4-6 membered heterocycloalkyl, eachof which is optionally substituted with 1, 2, 3, or 4 independentlyselected R³ groups. In some embodiments, Cy is selected from acyclopropyl ring, a phenyl ring, an azetidine ring, a pyrrolidine ring,a piperidine ring, 3-oxo-morpholin-6-yl, 2-oxo-pyrrolidin-4-yl,2-oxo-oxazolidin-4-yl, 2-oxo-oxazolidin-5-yl, a pyrazole ring, apyridine ring, and a pyrimidine ring, each of which is optionallysubstituted with 1, 2, 3, or 4 independently selected R³ groups.

In some embodiments:

each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo,CN, C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups;

each Cy¹ is independently C₃₋₇ cycloalkyl, which is optionallysubstituted with 1, 2, 3, or 4 independently selected R¹¹ groups;

each R^(a1), R^(c1), and R^(d1) is independently selected from H andC₁₋₆ alkyl, wherein said C₁₋₆ alkyl is optionally substituted with 1, 2,or 3 independently selected R¹¹ groups;

each R^(b1) is independently C₁₋₆ alkyl, which is optionally substitutedwith 1, 2, or 3 independently selected R¹¹ groups; and

each R¹¹ is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃haloalkoxy, amino, C₁₋₃ alkylamino, di(C₁₋₃ alkyl)amino, C₁₋₃alkylcarbonyl, C₁₋₃ alkoxycarbonyl, carbamyl, C₁₋₃ alkylcarbamyl, ordi(C₁₋₃ alkyl)carbamyl.

In some embodiments:

R² is C₁₋₃ alkyl or C₁₋₃ fluoroalkyl;

R⁴ is halo, CN, or C₁₋₃ alkyl;

R⁵ is halo, CN, or C₁₋₃ alkyl;

Cy is selected from C₃₋₆ cycloalkyl, 4-6 membered heterocycloalkyl,phenyl, and 5-6 membered heteroaryl, each of which is optionallysubstituted with 1, 2, 3, or 4 independently selected R³ group;

each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo,CN, C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups;

each Cy¹ is independently C₃₋₇ cycloalkyl, which is optionallysubstituted with 1, 2, 3, or 4 independently selected R¹¹ groups;

each R^(a1), R^(c1), and R^(d1) is independently selected from H andC₁₋₆ alkyl, wherein said C₁₋₆ alkyl is optionally substituted with 1, 2,or 3 independently selected R¹¹ groups;

each R^(b1) is independently C₁₋₆ alkyl, which is optionally substitutedwith 1, 2, or 3 independently selected R¹¹ groups; and

each R¹¹ is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃haloalkoxy, amino, C₁₋₃ alkylamino, di(C₁₋₃ alkyl)amino, C₁₋₃alkylcarbonyl, C₁₋₃ alkoxycarbonyl, carbamyl, C₁₋₃ alkylcarbamyl, ordi(C₁₋₃ alkyl)carbamyl.

In some embodiments:

R² is methyl, ethyl, or 2,2-difluoromethyl;

R⁴ is F, Cl, CN, or methyl;

R⁵ is Cl, CN, or methyl;

Cy is selected from C₃₋₆ cycloalkyl, 4-6 membered heterocycloalkyl,phenyl, and 5-6 membered heteroaryl, each of which is optionallysubstituted with 1, 2, 3, or 4 independently selected R³ group;

each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo,CN, C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups;

each Cy¹ is independently C₃₋₇ cycloalkyl, which is optionallysubstituted with 1, 2, 3, or 4 independently selected R¹¹ groups;

each R^(a1), R^(c1), and R^(d1) is independently selected from H andC₁₋₆ alkyl, wherein said C₁₋₆ alkyl is optionally substituted with 1, 2,or 3 independently selected R¹¹ groups;

each R^(b1) is independently C₁₋₆ alkyl, which is optionally substitutedwith 1, 2, or 3 independently selected R¹¹ groups; and

each R¹¹ is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃haloalkoxy, amino, C₁₋₃ alkylamino, di(C₁₋₃ alkyl)amino, C₁₋₃alkylcarbonyl, C₁₋₃ alkoxycarbonyl, carbamyl, C₁₋₃ alkylcarbamyl, ordi(C₁₋₃ alkyl)carbamyl.

In some embodiments:

R² is methyl, ethyl, or 2,2-difluoromethyl;

R⁴ is F, Cl, CN, or methyl;

R⁵ is Cl, CN, or methyl;

Cy is selected from a cyclopropyl ring, a phenyl ring, an azetidinering, a pyrrolidine ring, a piperidine ring, 3-oxo-morpholin-6-yl,2-oxo-pyrrolidin-4-yl, 2-oxo-oxazolidin-4-yl, 2-oxo-oxazolidin-5-yl, apyrazole ring, a pyridine ring, and a pyrimidine ring, each of which isoptionally substituted with 1, 2, 3, or 4 independently selected R³groups.

each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo,C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups;

each Cy¹ is independently selected from cyclopropyl and cyclobutyl, eachof which is optionally substituted with 1, 2, 3, or 4 independentlyselected R¹¹ groups;

each R^(a1), R^(c1), and R^(d1) is independently selected from H andC₁₋₄ alkyl; wherein said C₁₋₄ alkyl is optionally substituted with 1, 2,or 3 independently selected R¹¹ groups;

each R^(b1) is independently C₁₋₄ alkyl, which is optionally substitutedwith 1, 2, or 3 independently selected R¹¹ groups;

each R¹¹ is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃alkoxy-C₁₋₃ alkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkoxy, amino, C₁₋₃ alkylamino,di(C₁₋₃ alkyl)amino, C₁₋₃ alkylcarbonyl, C₁₋₄ alkoxycarbonyl, carbamyl,C₁₋₃ alkylcarbamyl, or di(C₁₋₃ alkyl)carbamyl.

In some embodiments, the compound is a compound of Formula II:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound is a compound of Formula III:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound is a compound of Formula IV:

or a pharmaceutically acceptable salt thereof, wherein:

G is NH, n is 1, and V is O; or

G is NH, n is 0, and V is O or CH₂; or

G is O, n is 0 and V is NH.

In some embodiments, the compound is a compound of Formula IVa:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound is a compound of Formula IVb:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound is a compound of Formula IVc:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound is a compound of Formula IVd:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the starred carbon in Formula I:

is a chiral carbon and said compound or said salt is the (S)-enantiomer.

In some embodiments, the compounds are those described in U.S. patentapplication Ser. No. 13/601,349, filed Aug. 31, 2012 (US Patent Publ.No. 2013/0059835), which is incorporated herein by reference in itsentirety.

In some embodiments, the method is a method of treating idiopathicthrombocytopenic purpura (or idiopathic immune thrombocytopenic purpura)(ITP). In some embodiments, the ITP is relapsed ITP. In someembodiments, the ITP is refractory ITP.

In some embodiments, the method is a method of treating autoimmunehemolytic anemia (AIHA).

In some embodiments, the method is a method is a method of treatingvasculitis. In some embodiments, the vasculitis is Behcet's disease,Cogan's syndrome, giant cell arteritis, polymyalgia rheumatica (PMR),Takayasu's arteritis, Buerger's disease (thromboangiitis obliterans),central nervous system vasculitis, Kawasaki disease, polyarteritisnodosa, Churg-Strauss syndrome, mixed cryoglobulinemia vasculitis(essential or hepatitis C virus (HCV)-induced), Henoch-Schonlein purpura(HSP), hypersensitivity vasculitis, microscopic polyangiitis, Wegener'sgranulomatosis, or anti-neutrophil cytoplasm antibody associated (ANCA)systemic vasculitis (AASV). In some embodiments, the method is a methodof treating nephritis.

In some embodiments, the method of treating non-Hodgkin lymphoma (NHL)is relapsed or refractory NHL or recucurrent follicular NHL.

In som embodiments, the present application provides a method oftreating an aggressive lymphoma (e.g., germinal center B cell-like (GCB)or activated B cell-like (ABC)) in a patient, comprising administering atherapeutic amount of any of the compounds described herein to saidpatient, or a pharmaceutically acceptable salt thereof.

In some embodiments, the present application provides a method oftreating acute myeloid leukemia in a patient, comprising administering atherapeutic amount of any of the compounds described herein to saidpatient, or a pharmaceutically acceptable salt thereof.

In some embodiments, the present application provides a method oftreating Burkitt lymphoma in a patient, comprising administering atherapeutic amount of any of the compounds described herein to saidpatient, or a pharmaceutically acceptable salt thereof.

It is appreciated that certain features of the invention, which are, forclarity, described in the context of separate embodiments, can also beprovided in combination in a single embodiment. Conversely, variousfeatures of the invention which are, for brevity, described in thecontext of a single embodiment, can also be provided separately or inany suitable subcombination.

At various places in the present specification, divalent linkingsubstituents are described. It is specifically intended that eachdivalent linking substituent include both the forward and backward formsof the linking substituent. For example, —NR(CR′R″)_(n)— includes both—NR(CR′R″)_(n)— and —(CR′R″)_(n)NR—. Where the structure clearlyrequires a linking group, the Markush variables listed for that groupare understood to be linking groups.

The term “n-membered” where n is an integer typically describes thenumber of ring-forming atoms in a moiety where the number ofring-forming atoms is n. For example, piperidinyl is an example of a6-membered heterocycloalkyl ring, pyrazolyl is an example of a5-membered heteroaryl ring, pyridyl is an example of a 6-memberedheteroaryl ring, and 1,2,3,4-tetrahydro-naphthalene is an example of a10-membered cycloalkyl group.

As used herein, the phrase “optionally substituted” means unsubstitutedor substituted. As used herein, the term “substituted” means that ahydrogen atom is removed and replaced by a substituent. It is to beunderstood that substitution at a given atom is limited by valency.

Throughout the definitions, the term “C_(n-m)” indicates a range whichincludes the endpoints, wherein n and m are integers and indicate thenumber of carbons. Examples include C₁₋₄, C₁₋₆, and the like.

As used herein, the term “C_(n-m) alkyl”, employed alone or incombination with other terms, refers to a saturated hydrocarbon groupthat may be straight-chain or branched, having n to m carbons. In someembodiments, the alkyl group contains from 1 to 6 carbon atoms, from 1to 4 carbon atoms, from 1 to 3 carbon atoms, or 1 to 2 carbon atoms.Examples of alkyl moieties include, but are not limited to, chemicalgroups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl,isobutyl, sec-butyl; higher homologs such as 2-methyl-1-butyl, n-pentyl,3-pentyl, n-hexyl, 1,2,2-trimethylpropyl, and the like.

As used herein, “C_(n-m) alkenyl” refers to an alkyl group having one ormore double carbon-carbon bonds and having n to m carbons. In someembodiments, the alkenyl moiety contains 2 to 6, 2 to 4, or 2 to 3carbon atoms. Example alkenyl groups include, but are not limited to,ethenyl, n-propenyl, isopropenyl, n-butenyl, sec-butenyl, and the like.

As used herein, “C_(n-m) alkynyl” refers to an alkyl group having one ormore triple carbon-carbon bonds and having n to m carbons. Examplealkynyl groups include, but are not limited to, ethynyl, propyn-1-yl,propyn-2-yl, and the like. In some embodiments, the alkynyl moietycontains 2 to 6, 2 to 4, or 2 to 3 carbon atoms.

As used herein, the term “alkylene”, employed alone or in combinationwith other terms, refers to a divalent alkyl linking group. Examples ofalkylene groups include, but are not limited to, ethan-1,2-diyl,propan-1,3-diyl, propan-1,2-diyl, butan-1,4-diyl, butan-1,3-diyl,butan-1,2-diyl, 2-methyl-propan-1,3-diyl, and the like.

As used herein, the term “C_(n-m) alkoxy”, employed alone or incombination with other terms, refers to a group of formula —O-alkyl,wherein the alkyl group has n to m carbons. Example alkoxy groupsinclude methoxy, ethoxy, propoxy (e.g., n-propoxy and isopropoxy),t-butoxy, and the like. In some embodiments, the alkyl group has 1 to 6,1 to 4, or 1 to 3 carbon atoms.

As used herein, the term “C_(n-m) alkylamino” refers to a group offormula —NH(alkyl), wherein the alkyl group has n to m carbon atoms. Insome embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbonatoms.

As used herein, the term “C_(n-m) alkoxycarbonyl” refers to a group offormula —C(O)O— alkyl, wherein the alkyl group has n to m carbon atoms.In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3carbon atoms.

As used herein, the term “C_(n-m) alkylcarbonyl” refers to a group offormula —C(O)-alkyl, wherein the alkyl group has n to m carbon atoms. Insome embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbonatoms.

As used herein, the term “C_(n-m) alkylcarbonylamino” refers to a groupof formula —NHC(O)-alkyl, wherein the alkyl group has n to m carbonatoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to3 carbon atoms.

As used herein, the term “C_(n-m) alkylsulfonylamino” refers to a groupof formula —NHS(O)₂-alkyl, wherein the alkyl group has n to m carbonatoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to3 carbon atoms.

As used herein, the term “aminosulfonyl” refers to a group of formula—S(O)₂NH₂.

As used herein, the term “C_(n-m) alkylaminosulfonyl” refers to a groupof formula —S(O)₂NH(alkyl), wherein the alkyl group has n to m carbonatoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to3 carbon atoms.

As used herein, the term “di(C_(n-m) alkyl)aminosulfonyl” refers to agroup of formula —S(O)₂N(alkyl)₂, wherein each alkyl group independentlyhas n to m carbon atoms. In some embodiments, each alkyl group has,independently, 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, the term “aminosulfonylamino” refers to a group offormula —NHS(O)₂NH₂.

As used herein, the term “C_(n-m) alkylaminosulfonylamino” refers to agroup of formula —NHS(O)₂NH(alkyl), wherein the alkyl group has n to mcarbon atoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4,or 1 to 3 carbon atoms.

As used herein, the term “di(C_(n-m) alkyl)aminosulfonylamino” refers toa group of formula —NHS(O)₂N(alkyl)₂, wherein each alkyl groupindependently has n to m carbon atoms. In some embodiments, each alkylgroup has, independently, 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, the term “aminocarbonylamino”, employed alone or incombination with other terms, refers to a group of formula —NHC(O)NH₂.

As used herein, the term “C_(n-m) alkylaminocarbonylamino” refers to agroup of formula —NHC(O)NH(alkyl), wherein the alkyl group has n to mcarbon atoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4,or 1 to 3 carbon atoms.

As used herein, the term “di(C_(n-m) alkyl)aminocarbonylamino” refers toa group of formula —NHC(O)N(alkyl)₂, wherein each alkyl groupindependently has n to m carbon atoms. In some embodiments, each alkylgroup has, independently, 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, the term “C_(n-m) alkylcarbamyl” refers to a group offormula —C(O)—NH(alkyl), wherein the alkyl group has n to m carbonatoms. In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to3 carbon atoms.

As used herein, the term “thio” refers to a group of formula —SH.

As used herein, the term “C_(n-m) alkylthio” refers to a group offormula —S-alkyl, wherein the alkyl group has n to m carbon atoms. Insome embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbonatoms.

As used herein, the term “C_(n-m) alkylsulfinyl” refers to a group offormula —S(O)-alkyl, wherein the alkyl group has n to m carbon atoms. Insome embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbonatoms.

As used herein, the term “C_(n-m) alkylsulfonyl” refers to a group offormula —S(O)₂-alkyl, wherein the alkyl group has n to m carbon atoms.In some embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3carbon atoms.

As used herein, the term “amino” refers to a group of formula —NH₂.

As used herein, the term “carbamyl” to a group of formula —C(O)NH₂.

As used herein, the term “carbonyl”, employed alone or in combinationwith other terms, refers to a —C(O)— group.

As used herein, the term “cyano-C₁₋₃ alkyl” refers to a group of formula—(C₁₋₃ alkylene)-CN.

As used herein, the term “HO—C₁₋₃ alkyl” refers to a group of formula—(C₁₋₃ alkylene)-OH.

As used herein, the term “C₁₋₃ alkoxy-C₁₋₃ alkyl” refers to a group offormula —(C₁₋₃ alkylene)-O(C₁₋₃ alkyl).

As used herein, the term “carboxy” refers to a group of formula —C(O)OH.

As used herein, the term “di(C_(n-m)-alkyl)amino” refers to a group offormula —N(alkyl)₂, wherein the two alkyl groups each has,independently, n to m carbon atoms. In some embodiments, each alkylgroup independently has 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, the term “di(C_(n-m)-alkyl)carbamyl” refers to a groupof formula —C(O)N(alkyl)₂, wherein the two alkyl groups each has,independently, n to m carbon atoms. In some embodiments, each alkylgroup independently has 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, “halo” refers to F, Cl, Br, or I. In some embodiments,the halo group is F or Cl.

As used herein, “C_(n-m) haloalkoxy” refers to a group of formula—O-haloalkyl having n to m carbon atoms. An example haloalkoxy group isOCF₃. In some embodiments, the haloalkoxy group is fluorinated only. Insome embodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbonatoms.

As used herein, the term “C_(n-m) haloalkyl”, employed alone or incombination with other terms, refers to an alkyl group having from onehalogen atom to 2s+1 halogen atoms which may be the same or different,where “s” is the number of carbon atoms in the alkyl group, wherein thealkyl group has n to m carbon atoms. In some embodiments, the haloalkylgroup is fluorinated only (e.g., a “fluoroalkyl” group). In someembodiments, the alkyl group has 1 to 6, 1 to 4, or 1 to 3 carbon atoms.

As used herein, “cycloalkyl” refers to non-aromatic cyclic hydrocarbonsincluding cyclized alkyl and/or alkenyl groups. Cycloalkyl groups canhave 3, 4, 5, 6, or 7 ring-forming carbons (C₃₋₇). Ring-forming carbonatoms of a cycloalkyl group can be optionally substituted by oxo orsulfido. Cycloalkyl groups also include cycloalkylidenes. Examplecycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, cycloheptyl, cyclopentenyl, cyclohexenyl, cyclohexadienyl,cycloheptatrienyl, and the like. In some embodiments, cycloalkyl iscyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. Also included inthe definition of cycloalkyl are moieties that have one or more aromaticrings fused (i.e., having a bond in common with) to the cycloalkyl ring,for example, benzo or thienyl derivatives of cyclopentane, cyclohexane,and the like.

As used herein, “heteroaryl” refers to a monocyclic aromatic heterocyclehaving at least one heteroatom ring member selected from sulfur, oxygen,and nitrogen. In some embodiments, the heteroaryl ring has 1, 2, 3, or 4heteroatom ring members independently selected from nitrogen, sulfur andoxygen. In some embodiments, any ring-forming N in a heteroaryl moietycan be an N-oxide. In some embodiments, the heteroaryl has 5-6 ringatoms and 1 or 2 heteroatom ring members independently selected fromnitrogen, sulfur and oxygen. In some embodiments, the heteroaryl is afive-membered or six-membered heteroaryl ring.

A five-membered heteroaryl ring is a heteroaryl with a ring having fivering atoms wherein one or more (e.g., 1, 2, or 3) ring atoms areindependently selected from N, O, and S. Exemplary five-membered ringheteroaryls are thienyl, furyl, pyrrolyl, imidazolyl, thiazolyl,oxazolyl, pyrazolyl, isothiazolyl, isoxazolyl, 1,2,3-triazolyl,tetrazolyl, 1,2,3-thiadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-triazolyl,1,2,4-thiadiazolyl, 1,2,4-oxadiazolyl, 1,3,4-triazolyl,1,3,4-thiadiazolyl, and 1,3,4-oxadiazolyl.

A six-membered heteroaryl ring is a heteroaryl with a ring having sixring atoms wherein one or more (e.g., 1, 2, or 3) ring atoms areindependently selected from N, O, and S. Exemplary six-membered ringheteroaryls are pyridyl, pyrazinyl, pyrimidinyl, triazinyl andpyridazinyl.

As used herein, “heterocycloalkyl” refers to non-aromatic monocyclicheterocycleshaving one or more ring-forming heteroatoms selected from O,N, or S. Included in heterocycloalkyl are monocyclic 4-, 5-, 6-, and7-membered heterocycloalkyl groups. Example heterocycloalkyl groupsinclude pyrrolidin-2-one, 1,3-isoxazolidin-2-one, pyranyl,tetrahydropuran, oxetanyl, azetidinyl, morpholino, thiomorpholino,piperazinyl, tetrahydrofuranyl, tetrahydrothienyl, piperidinyl,pyrrolidinyl, isoxazolidinyl, isothiazolidinyl, pyrazolidinyl,oxazolidinyl, thiazolidinyl, imidazolidinyl, azepanyl,3-oxo-morpholin-6-yl, 2-oxo-pyrrolidin-4-yl, 2-oxo-oxazolidin-4-yl,2-oxo-oxazolidin-5-yl, and the like. Ring-forming carbon atoms andheteroatoms of a heterocycloalkyl group can be optionally substituted byoxo or sulfido (e.g., C(O), S(O), C(S), or S(O)₂, etc.). Theheterocycloalkyl group can be attached through a ring-forming carbonatom or a ring-forming heteroatom. In some embodiments, theheterocycloalkyl group contains 0 to 3 double bonds. In someembodiments, the heterocycloalkyl group contains 0 to 2 double bonds. Insome embodiments, the heterocycloalkyl has 4-7 or 4-6 ring atoms with 1or 2 heteroatoms independently selected from nitrogen, oxygen or sulfurand having one or more oxidized ring members.

At certain places, the definitions or embodiments refer to specificrings (e.g., an azetidine ring, a pyridine ring, etc.). Unless otherwiseindicated, these rings can be attached any ring member provided that thevalency of the atom is not exceeded. For example, an azetidine ring maybe attached at any position of the ring, whereas an azetidin-3-yl ringis attached at the 3-position.

The compounds described herein can be asymmetric (e.g., having one ormore stereocenters). All stereoisomers, such as enantiomers anddiastereomers, are intended unless otherwise indicated. Compounds of thepresent invention that contain asymmetrically substituted carbon atomscan be isolated in optically active or racemic forms. Methods on how toprepare optically active forms from optically inactive startingmaterials are known in the art, such as by resolution of racemicmixtures or by stereoselective synthesis. Many geometric isomers ofolefins, C═N double bonds, and the like can also be present in thecompounds described herein, and all such stable isomers are contemplatedin the present invention. Cis and trans geometric isomers of thecompounds of the present invention are described and may be isolated asa mixture of isomers or as separated isomeric forms.

In some embodiments, the compound has the (R)-configuration. In someembodiments, the compound has the (S)-configuration.

Resolution of racemic mixtures of compounds can be carried out by any ofnumerous methods known in the art. An example method includes fractionalrecrystallization using a chiral resolving acid which is an opticallyactive, salt-forming organic acid. Suitable resolving agents forfractional recrystallization methods are, for example, optically activeacids, such as the D and L forms of tartaric acid, diacetyltartaricacid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid orthe various optically active camphorsulfonic acids such asβ-camphorsulfonic acid.

Other resolving agents suitable for fractional crystallization methodsinclude stereoisomerically pure forms of α-methylbenzylamine (e.g., Sand R forms, or diastereomerically pure forms), 2-phenylglycinol,norephedrine, ephedrine, N-methylephedrine, cyclohexylethylamine,1,2-diaminocyclohexane, and the like.

Resolution of racemic mixtures can also be carried out by elution on acolumn packed with an optically active resolving agent (e.g.,dinitrobenzoylphenylglycine). Suitable elution solvent composition canbe determined by one skilled in the art.

Compounds described herein also include tautomeric forms. Tautomericforms result from the swapping of a single bond with an adjacent doublebond together with the concomitant migration of a proton. Tautomericforms include prototropic tautomers which are isomeric protonationstates having the same empirical formula and total charge. Exampleprototropic tautomers include ketone—enol pairs, amide—imidic acidpairs, lactam—lactim pairs, enamine-imine pairs, and annular forms wherea proton can occupy two or more positions of a heterocyclic system, forexample, 1H- and 3H-imidazole, 1H-, 2H- and 4H-1,2,4-triazole, 1H- and2H-isoindole, and 1H- and 2H-pyrazole. Tautomeric forms can be inequilibrium or sterically locked into one form by appropriatesubstitution.

Compounds described herein can also include all isotopes of atomsoccurring in the intermediates or final compounds. Isotopes includethose atoms having the same atomic number but different mass numbers.For example, isotopes of hydrogen include tritium and deuterium.

The term, “compound,” as used herein is meant to include allstereoisomers, geometric iosomers, tautomers, and isotopes of thestructures depicted. Compounds herein identified by name or structure asone particular tautomeric form are intended to include other tautomericforms unless otherwise specified.

All compounds, and pharmaceutically acceptable salts thereof, can befound together with other substances such as water and solvents (e.g.hydrates and solvates) or can be isolated.

In some embodiments, the compounds described herein, or salts thereof,are substantially isolated. By “substantially isolated” is meant thatthe compound is at least partially or substantially separated from theenvironment in which it was formed or detected. Partial separation caninclude, for example, a composition enriched in the compounds describedherein. Substantial separation can include compositions containing atleast about 50%, at least about 60%, at least about 70%, at least about80%, at least about 90%, at least about 95%, at least about 97%, or atleast about 99% by weight of the compounds described herein, or saltthereof. Methods for isolating compounds and their salts are routine inthe art.

The phrase “pharmaceutically acceptable” is employed herein to refer tothose compounds, materials, compositions, and/or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of human beings and animals without excessive toxicity,irritation, allergic response, or other problem or complication,commensurate with a reasonable benefit/risk ratio.

The expressions, “ambient temperature” and “room temperature” or “rt” asused herein, are understood in the art, and refer generally to atemperature, e.g. a reaction temperature, that is about the temperatureof the room in which the reaction is carried out, for example, atemperature from about 20° C. to about 30° C.

The present invention also includes pharmaceutically acceptable salts ofthe compounds described herein. As used herein, “pharmaceuticallyacceptable salts” refers to derivatives of the disclosed compoundswherein the parent compound is modified by converting an existing acidor base moiety to its salt form. Examples of pharmaceutically acceptablesalts include, but are not limited to, mineral or organic acid salts ofbasic residues such as amines; alkali or organic salts of acidicresidues such as carboxylic acids; and the like. The pharmaceuticallyacceptable salts of the present invention include the conventionalnon-toxic salts of the parent compound formed, for example, fromnon-toxic inorganic or organic acids. The pharmaceutically acceptablesalts of the present invention can be synthesized from the parentcompound which contains a basic or acidic moiety by conventionalchemical methods. Generally, such salts can be prepared by reacting thefree acid or base forms of these compounds with a stoichiometric amountof the appropriate base or acid in water or in an organic solvent, or ina mixture of the two; generally, non-aqueous media like ether, ethylacetate, alcohols (e.g., methanol, ethanol, iso-propanol, or butanol) oracetonitrile (ACN) are preferred. Lists of suitable salts are found inRemington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company,Easton, Pa., 1985, p. 1418 and Journal of Pharmaceutical Science, 66, 2(1977), each of which is incorporated herein by reference in itsentirety.

Methods

The compounds described herein can modulate activity of one or more ofvarious kinases including, for example, phosphoinositide 3-kinases(PI3Ks). The term “modulate” is meant to refer to an ability to increaseor decrease the activity of one or more members of the PI3K family.Accordingly, the compounds described herein can be used in methods ofmodulating a PI3K by contacting the PI3K with any one or more of thecompounds or compositions described herein. In some embodiments,compounds of the present invention can act as inhibitors of one or morePI3Ks. In further embodiments, the compounds described herein can beused to modulate activity of a PI3K in an individual in need ofmodulation of the receptor by administering a modulating amount of acompound described herein, or a pharmaceutically acceptable saltthereof. In some embodiments, modulating is inhibiting.

Given that cancer cell growth and survival is impacted by multiplesignaling pathways, the present invention is useful for treating diseasestates characterized by drug resistant kinase mutants. In addition,different kinase inhibitors, exhibiting different preferences in thekinases which they modulate the activities of, may be used incombination. This approach could prove highly efficient in treatingdisease states by targeting multiple signaling pathways, reduce thelikelihood of drug-resistance arising in a cell, and reduce the toxicityof treatments for disease.

Kinases to which the present compounds bind and/or modulate (e.g.,inhibit) include any member of the PI3K family. In some embodiments, thePI3K is PI3Kα, PI3Kβ, PI3Kγ, or PI3Kδ. In some embodiments, the PI3K isPI3Kγ or PI3Kδ. In some embodiments, the PI3K is PI3Kγ. In someembodiments, the PI3K is PI3Kδ. In some embodiments, the PI3K includes amutation. A mutation can be a replacement of one amino acid for another,or a deletion of one or more amino acids. In such embodiments, themutation can be present in the kinase domain of the PI3K.

In some embodiments, more than one compound described herein is used toinhibit the activity of one kinase (e.g., PI3Kγ or PI3Kδ).

In some embodiments, more than one compound described herein is used toinhibit more than one kinase, such as at least two kinases (e.g., PI3Kγand PI3Kδ).

In some embodiments, one or more of the compounds is used in combinationwith another kinase inhibitor to inhibit the activity of one kinase(e.g., PI3Kγ or PI3Kδ).

In some embodiments, one or more of the compounds is used in combinationwith another kinase inhibitor to inhibit the activities of more than onekinase (e.g., PI3Kγ or PI3Kδ), such as at least two kinases.

The compounds described herein can be selective. By “selective” is meantthat the compound binds to or inhibits a kinase with greater affinity orpotency, respectively, compared to at least one other kinase. In someembodiments, the compounds described herein are selective inhibitors ofPI3Kγ or PI3Kδ over PI3Kα and/or PI3Kβ. In some embodiments, thecompounds described herein are selective inhibitors of PI3Kδ (e.g., overPI3Kα, PI3Kβ and PI3Kγ). In some embodiments, the compounds describedherein are selective inhibitors of PI3Kγ (e.g., over PI3K, PI3Kβ andPI3Kδ). In some embodiments, selectivity can be at least about 2-fold,5-fold, 10-fold, at least about 20-fold, at least about 50-fold, atleast about 100-fold, at least about 200-fold, at least about 500-foldor at least about 1000-fold. Selectivity can be measured by methodsroutine in the art. In some embodiments, selectivity can be tested atthe K_(m) ATP concentration of each enzyme. In some embodiments, theselectivity of compounds described herein can be determined by cellularassays associated with particular PI3K kinase activity.

As used herein, the term “contacting” refers to the bringing together ofindicated moieties in an in vitro system or an in vivo system. Forexample, “contacting” a PI3K with a compound described herein includesthe administration of a compound of the present invention to anindividual or patient, such as a human, having a PI3K, as well as, forexample, introducing a compound described herein into a samplecontaining a cellular or purified preparation containing the PI3K.

As used herein, the term “individual” or “patient,” usedinterchangeably, refers to any animal, including mammals, preferablymice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep,horses, or primates, and most preferably humans.

As used herein, the phrase “therapeutically effective amount” refers tothe amount of active compound or pharmaceutical agent that elicits thebiological or medicinal response that is being sought in a tissue,system, animal, individual or human by a researcher, veterinarian,medical doctor or other clinician. In some embodiments, the dosage ofthe compound, or a pharmaceutically acceptable salt thereof,administered to a patient or individual is about 1 mg to about 2 g,about 1 mg to about 1000 mg, about 1 mg to about 500 mg, about 1 mg toabout 100 mg, about 1 mg to 50 mg, or about 50 mg to about 500 mg.

As used herein, the term “treating” or “treatment” refers to one or moreof (1) preventing the disease; for example, preventing a disease,condition or disorder in an individual who may be predisposed to thedisease, condition or disorder but does not yet experience or displaythe pathology or symptomatology of the disease; (2) inhibiting thedisease; for example, inhibiting a disease, condition or disorder in anindividual who is experiencing or displaying the pathology orsymptomatology of the disease, condition or disorder (i.e., arrestingfurther development of the pathology and/or symptomatology); and (3)ameliorating the disease; for example, ameliorating a disease, conditionor disorder in an individual who is experiencing or displaying thepathology or symptomatology of the disease, condition or disorder (i.e.,reversing the pathology and/or symptomatology) such as decreasing theseverity of disease.

Combination Therapies

One or more additional pharmaceutical agents such as, for example,chemotherapeutics, anti-inflammatory agents, steroids,immunosuppressants, as well as Bcr-Abl, Flt-3, EGFR, HER2, JAK (e.g.,JAK1 or JAK2), c-MET, VEGFR, PDGFR, cKit, IGF-1R, RAF, FAK, Akt mTOR,PIM, and AKT (e.g., AKT1, AKT2, or AKT3) kinase inhibitors such as, forexample, those described in WO 2006/056399, or other agents such as,therapeutic antibodies can be used in combination with the compounds ofthe present invention for treatment of PI3K-associated diseases,disorders or conditions. The one or more additional pharmaceuticalagents can be administered to a patient simultaneously or sequentially.

Example antibodies for use in combination therapy include but are notlimited to Trastuzumab (e.g. anti-HER2), Ranibizumab (e.g. anti-VEGF-A),Bevacizumab (trade name Avastin, e.g. anti-VEGF, Panitumumab (e.g.anti-EGFR), Cetuximab (e.g. anti-EGFR), Rituxan (anti-CD20) andantibodies directed to c-MET.

One or more of the following agents may be used in combination with thecompounds of the present invention and are presented as a non limitinglist: a cytostatic agent, cisplatin, doxorubicin, taxotere, taxol,etoposide, irinotecan, camptostar, topotecan, paclitaxel, docetaxel,epothilones, tamoxifen, 5-fluorouracil, methoxtrexate, temozolomide,cyclophosphamide, SCH 66336, R115777, L778,123, BMS 214662, Iressa,Tarceva, antibodies to EGFR, Gleevec™ intron, ara-C, adriamycin,cytoxan, gemcitabine, Uracil mustard, Chlormethine, Ifosfamide,Melphalan, Chlorambucil, Pipobroman, Triethylenemelamine,Triethylenethiophosphoramine, Busulfan, Carmustine, Lomustine,Streptozocin, Dacarbazine, Floxuridine, Cytarabine, 6-Mercaptopurine,6-Thioguanine, Fludarabine phosphate, oxaliplatin, leucovirin, ELOXATIN™Pentostatine, Vinblastine, Vincristine, Vindesine, Bleomycin,Dactinomycin, Daunorubicin, Doxorubicin, Epirubicin, Idarubicin,Mithramycin, Deoxycoformycin, Mitomycin-C, L-Asparaginase, Teniposide17.alpha.-Ethinylestradiol, Diethylstilbestrol, Testosterone,Prednisone, Fluoxymesterone, Dromostanolone propionate, Testolactone,Megestrolacetate, Methylprednisolone, Methyltestosterone, Prednisolone,Triamcinolone, Chlorotrianisene, Hydroxyprogesterone, Aminoglutethimide,Estramustine, Medroxyprogesteroneacetate, Leuprolide, Flutamide,Toremifene, goserelin, Cisplatin, Carboplatin, Hydroxyurea, Amsacrine,Procarbazine, Mitotane, Mitoxantrone, Levamisole, Navelbene,Anastrazole, Letrazole, Capecitabine, Reloxafine, Droloxafine,Hexamethylmelamine, Avastin, herceptin, Bexxar, Velcade, Zevalin,Trisenox, Xeloda, Vinorelbine, Porfimer, Erbitux, Liposomal, Thiotepa,Altretamine, Melphalan, Trastuzumab, Lerozole, Fulvestrant, Exemestane,Fulvestrant, Ifosfomide, Rituximab, C225, Campath, Clofarabine,cladribine, aphidicolon, rituxan, sunitinib, dasatinib, tezacitabine,Smll, fludarabine, pentostatin, triapine, didox, trimidox, amidox, 3-AP,MDL-101,731, bendamustine (Treanda), ofatumumab, or GS-1101 (also knownas CAL-101).

Example chemotherapeutics include proteosome inhibitors (e.g.,bortezomib), thalidomide, revlimid, and DNA-damaging agents such asmelphalan, doxorubicin, cyclophosphamide, vincristine, etoposide,carmustine, and the like.

Example steroids include coriticosteroids such as dexamethasone orprednisone.

Example Bcr-Abl inhibitors include the compounds, and pharmaceuticallyacceptable salts thereof, of the genera and species disclosed in U.S.Pat. No. 5,521,184, WO 04/005281, and U.S. Ser. No. 60/578,491.

Example suitable Flt-3 inhibitors include compounds, and theirpharmaceutically acceptable salts, as disclosed in WO 03/037347, WO03/099771, and WO 04/046120.

Example suitable RAF inhibitors include compounds, and theirpharmaceutically acceptable salts, as disclosed in WO 00/09495 and WO05/028444.

Example suitable FAK inhibitors include compounds, and theirpharmaceutically acceptable salts, as disclosed in WO 04/080980, WO04/056786, WO 03/024967, WO 01/064655, WO 00/053595, and WO 01/014402.

Example suitable mTOR inhibitors include compounds, and theirpharmaceutically acceptable salts, as disclosed in WO 2011/025889.

In some embodiments, the compounds of the invention can be used incombination with one or more other kinase inhibitors including imatinib,particularly for treating patients resistant to imatinib or other kinaseinhibitors.

In some embodiments, the compounds of the invention can be used incombination with a chemotherapeutic in the treatment of cancer, such asmultiple myeloma, and may improve the treatment response as compared tothe response to the chemotherapeutic agent alone, without exacerbationof its toxic effects. Examples of additional pharmaceutical agents usedin the treatment of multiple myeloma, for example, can include, withoutlimitation, melphalan, melphalan plus prednisone [MP], doxorubicin,dexamethasone, and Velcade (bortezomib). Further additional agents usedin the treatment of multiple myeloma include Bcr-Abl, Flt-3, RAF and FAKkinase inhibitors. Additive or synergistic effects are desirableoutcomes of combining a PI3K inhibitor of the present invention with anadditional agent. Furthermore, resistance of multiple myeloma cells toagents such as dexamethasone may be reversible upon treatment with thePI3K inhibitor of the present invention. The agents can be combined withthe present compound in a single or continuous dosage form, or theagents can be administered simultaneously or sequentially as separatedosage forms.

In some embodiments, a corticosteroid such as dexamethasone isadministered to a patient in combination with the compounds of theinvention where the dexamethasone is administered intermittently asopposed to continuously.

In some further embodiments, combinations of the compounds of theinvention with other therapeutic agents can be administered to a patientprior to, during, and/or after a bone marrow transplant or stem celltransplant.

Pharmaceutical Formulations and Dosage Forms

When employed as pharmaceuticals, the compounds described herein can beadministered in the form of pharmaceutical compositions. Thesecompositions can be prepared in a manner well known in thepharmaceutical art, and can be administered by a variety of routes,depending upon whether local or systemic treatment is desired and uponthe area to be treated.

Administration may be topical (including transdermal, epidermal,ophthalmic and to mucous membranes including intranasal, vaginal andrectal delivery), pulmonary (e.g., by inhalation or insufflation ofpowders or aerosols, including by nebulizer; intratracheal orintranasal), oral or parenteral. Parenteral administration includesintravenous, intraarterial, subcutaneous, intraperitoneal intramuscularor injection or infusion; or intracranial, e.g., intrathecal orintraventricular, administration. Parenteral administration can be inthe form of a single bolus dose, or may be, for example, by a continuousperfusion pump. Pharmaceutical compositions and formulations for topicaladministration may include transdermal patches, ointments, lotions,creams, gels, drops, suppositories, sprays, liquids and powders.Conventional pharmaceutical carriers, aqueous, powder or oily bases,thickeners and the like may be necessary or desirable.

This invention also includes pharmaceutical compositions which contain,as the active ingredient, the compound described herein or apharmaceutically acceptable salt thereof, in combination with one ormore pharmaceutically acceptable carriers (excipients). In someembodiments, the composition is suitable for topical administration. Inmaking the compositions of the invention, the active ingredient istypically mixed with an excipient, diluted by an excipient or enclosedwithin such a carrier in the form of, for example, a capsule, sachet,paper, or other container. When the excipient serves as a diluent, itcan be a solid, semi-solid, or liquid material, which acts as a vehicle,carrier or medium for the active ingredient. Thus, the compositions canbe in the form of tablets, pills, powders, lozenges, sachets, cachets,elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solidor in a liquid medium), ointments containing, for example, up to 10% byweight of the active compound, soft and hard gelatin capsules,suppositories, sterile injectable solutions, and sterile packagedpowders.

In preparing a formulation, the active compound can be milled to providethe appropriate particle size prior to combining with the otheringredients. If the active compound is substantially insoluble, it canbe milled to a particle size of less than 200 mesh. If the activecompound is substantially water soluble, the particle size can beadjusted by milling to provide a substantially uniform distribution inthe formulation, e.g. about 40 mesh.

The compounds described herein may be milled using known millingprocedures such as wet milling to obtain a particle size appropriate fortablet formation and for other formulation types. Finely divided(nanoparticulate) preparations of the compounds described herein can beprepared by processes known in the art, e.g., see International App. No.WO 2002/000196.

Some examples of suitable excipients include lactose, dextrose, sucrose,sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates,tragacanth, gelatin, calcium silicate, microcrystalline cellulose,polyvinylpyrrolidone, cellulose, water, syrup, and methyl cellulose. Theformulations can additionally include: lubricating agents such as talc,magnesium stearate, and mineral oil; wetting agents; emulsifying andsuspending agents; preserving agents such as methyl- andpropylhydroxy-benzoates; sweetening agents; and flavoring agents. Thecompositions of the invention can be formulated so as to provide quick,sustained or delayed release of the active ingredient afteradministration to the patient by employing procedures known in the art.

The compositions can be formulated in a unit dosage form, each dosagecontaining from about 5 to about 1000 mg (1 g), more usually about 100to about 500 mg, of the active ingredient. The term “unit dosage forms”refers to physically discrete units suitable as unitary dosages forhuman subjects and other mammals, each unit containing a predeterminedquantity of active material calculated to produce the desiredtherapeutic effect, in association with a suitable pharmaceuticalexcipient.

In some embodiments, the compositions of the invention contain fromabout 5 to about 50 mg of the active ingredient. One having ordinaryskill in the art will appreciate that this embodies compositionscontaining about 5 to about 10, about 10 to about 15, about 15 to about20, about 20 to about 25, about 25 to about 30, about 30 to about 35,about 35 to about 40, about 40 to about 45, or about 45 to about 50 mgof the active ingredient.

In some embodiments, the compositions of the invention contain fromabout 50 to about 500 mg of the active ingredient. One having ordinaryskill in the art will appreciate that this embodies compositionscontaining about 50 to about 100, about 100 to about 150, about 150 toabout 200, about 200 to about 250, about 250 to about 300, about 350 toabout 400, or about 450 to about 500 mg of the active ingredient.

In some embodiments, the compositions of the invention contain fromabout 500 to about 1000 mg of the active ingredient. One having ordinaryskill in the art will appreciate that this embodies compositionscontaining about 500 to about 550, about 550 to about 600, about 600 toabout 650, about 650 to about 700, about 700 to about 750, about 750 toabout 800, about 800 to about 850, about 850 to about 900, about 900 toabout 950, or about 950 to about 1000 mg of the active ingredient.

Similar dosages may be used of the compounds described herein in themethods and uses of the invention.

The active compound can be effective over a wide dosage range and isgenerally administered in a pharmaceutically effective amount. It willbe understood, however, that the amount of the compound actuallyadministered will usually be determined by a physician, according to therelevant circumstances, including the condition to be treated, thechosen route of administration, the actual compound administered, theage, weight, and response of the individual patient, the severity of thepatient's symptoms, and the like.

For preparing solid compositions such as tablets, the principal activeingredient is mixed with a pharmaceutical excipient to form a solidpreformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, the active ingredient istypically dispersed evenly throughout the composition so that thecomposition can be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules. This solid preformulation isthen subdivided into unit dosage forms of the type described abovecontaining from, for example, about 0.1 to about 1000 mg of the activeingredient of the present invention.

The tablets or pills of the present invention can be coated or otherwisecompounded to provide a dosage form affording the advantage of prolongedaction. For example, the tablet or pill can comprise an inner dosage andan outer dosage component, the latter being in the form of an envelopeover the former. The two components can be separated by an enteric layerwhich serves to resist disintegration in the stomach and permit theinner component to pass intact into the duodenum or to be delayed inrelease. A variety of materials can be used for such enteric layers orcoatings, such materials including a number of polymeric acids andmixtures of polymeric acids with such materials as shellac, cetylalcohol, and cellulose acetate.

The liquid forms in which the compounds and compositions of the presentinvention can be incorporated for administration orally or by injectioninclude aqueous solutions, suitably flavored syrups, aqueous or oilsuspensions, and flavored emulsions with edible oils such as cottonseedoil, sesame oil, coconut oil, or peanut oil, as well as elixirs andsimilar pharmaceutical vehicles.

Compositions for inhalation or insufflation include solutions andsuspensions in pharmaceutically acceptable, aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedsupra. In some embodiments, the compositions are administered by theoral or nasal respiratory route for local or systemic effect.Compositions can be nebulized by use of inert gases. Nebulized solutionsmay be breathed directly from the nebulizing device or the nebulizingdevice can be attached to a face mask, tent, or intermittent positivepressure breathing machine. Solution, suspension, or powder compositionscan be administered orally or nasally from devices which deliver theformulation in an appropriate manner.

Topical formulations can contain one or more conventional carriers. Insome embodiments, ointments can contain water and one or morehydrophobic carriers selected from, for example, liquid paraffin,polyoxyethylene alkyl ether, propylene glycol, white Vaseline, and thelike. Carrier compositions of creams can be based on water incombination with glycerol and one or more other components, e.g.glycerinemonostearate, PEG-glycerinemonostearate and cetylstearylalcohol. Gels can be formulated using isopropyl alcohol and water,suitably in combination with other components such as, for example,glycerol, hydroxyethyl cellulose, and the like. In some embodiments,topical formulations contain at least about 0.1, at least about 0.25, atleast about 0.5, at least about 1, at least about 2, or at least about 5wt % of the compound described herein. The topical formulations can besuitably packaged in tubes of, for example, 100 g which are optionallyassociated with instructions for the treatment of the select indication,e.g., psoriasis or other skin condition.

The amount of compound or composition administered to a patient willvary depending upon what is being administered, the purpose of theadministration, such as prophylaxis or therapy, the state of thepatient, the manner of administration, and the like. In therapeuticapplications, compositions can be administered to a patient alreadysuffering from a disease in an amount sufficient to cure or at leastpartially arrest the symptoms of the disease and its complications.Effective doses will depend on the disease condition being treated aswell as by the judgment of the attending clinician depending uponfactors such as the severity of the disease, the age, weight and generalcondition of the patient, and the like.

The compositions administered to a patient can be in the form ofpharmaceutical compositions described above. These compositions can besterilized by conventional sterilization techniques, or may be sterilefiltered. Aqueous solutions can be packaged for use as is, orlyophilized, the lyophilized preparation being combined with a sterileaqueous carrier prior to administration. The pH of the compoundpreparations typically will be between 3 and 11, more preferably from 5to 9 and most preferably from 7 to 8. It will be understood that use ofcertain of the foregoing excipients, carriers, or stabilizers willresult in the formation of pharmaceutical salts.

The therapeutic dosage of a compound of the present invention can varyaccording to, for example, the particular use for which the treatment ismade, the manner of administration of the compound, the health andcondition of the patient, and the judgment of the prescribing physician.The proportion or concentration of a compound described herein in apharmaceutical composition can vary depending upon a number of factorsincluding dosage, chemical characteristics (e.g., hydrophobicity), andthe route of administration. For example, the compounds described hereincan be provided in an aqueous physiological buffer solution containingabout 0.1 to about 10% w/v of the compound for parenteraladministration. Some typical dose ranges are from about 1 rig/kg toabout 1 g/kg of body weight per day. In some embodiments, the dose rangeis from about 0.01 mg/kg to about 100 mg/kg of body weight per day. Thedosage is likely to depend on such variables as the type and extent ofprogression of the disease or disorder, the overall health status of theparticular patient, the relative biological efficacy of the compoundselected, formulation of the excipient, and its route of administration.Effective doses can be extrapolated from dose-response curves derivedfrom in vitro or animal model test systems.

The compositions of the invention can further include one or moreadditional pharmaceutical agents such as a chemotherapeutic, steroid,anti-inflammatory compound, or immunosuppressant, examples of which arelisted herein.

Kits

The present invention also includes pharmaceutical kits useful, forexample, in the treatment or prevention of PI3K-associated diseases ordisorders, such as cancer, which include one or more containerscontaining a pharmaceutical composition comprising a therapeuticallyeffective amount of a compound described herein. Such kits can furtherinclude, if desired, one or more of various conventional pharmaceuticalkit components, such as, for example, containers with one or morepharmaceutically acceptable carriers, additional containers, etc., aswill be readily apparent to those skilled in the art. Instructions,either as inserts or as labels, indicating quantities of the componentsto be administered, guidelines for administration, and/or guidelines formixing the components, can also be included in the kit.

Synthesis

Compounds described herein, including salts thereof, can be preparedusing known organic synthesis techniques and can be synthesizedaccording to any of numerous possible synthetic routes. In someembodiments, the compounds can be prepared as described in U.S. patentapplication Ser. No. 13/601,349, filed Aug. 31, 2012, which isincorporated herein by reference in its entirety.

The reactions for preparing compounds described herein can be carriedout in suitable solvents which can be readily selected by one of skillin the art of organic synthesis. Suitable solvents can be substantiallynon-reactive with the starting materials (reactants), the intermediates,or products at the temperatures at which the reactions are carried out,e.g., temperatures which can range from the solvent's freezingtemperature to the solvent's boiling temperature. A given reaction canbe carried out in one solvent or a mixture of more than one solvent.Depending on the particular reaction step, suitable solvents for aparticular reaction step can be selected by the skilled artisan.

Preparation of compounds described herein can involve the protection anddeprotection of various chemical groups. The need for protection anddeprotection, and the selection of appropriate protecting groups, can bereadily determined by one skilled in the art. The chemistry ofprotecting groups can be found, for example, in T. W. Greene and P. G.M. Wuts, Protective Groups in Organic Synthesis, 3^(rd) Ed., Wiley &Sons, Inc., New York (1999), which is incorporated herein by referencein its entirety.

Reactions can be monitored according to any suitable method known in theart. For example, product formation can be monitored by spectroscopicmeans, such as nuclear magnetic resonance spectroscopy (e.g., ¹H or¹³C), infrared spectroscopy, spectrophotometry (e.g., UV-visible), massspectrometry, or by chromatographic methods such as high performanceliquid chromatography (HPLC), liquid chromatography-mass spectroscopy(LCMS), or thin layer chromatography (TLC). Compounds can be purified bythose skilled in the art by a variety of methods, including highperformance liquid chromatography (HPLC) (“Preparative LC-MSPurification: Improved Compound Specific Method Optimization” Karl F.Blom, Brian Glass, Richard Sparks, Andrew P. Combs J. Combi. Chem. 2004,6(6), 874-883, which is incorporated herein by reference in itsentirety) and normal phase silica chromatography.

For example, compounds of Formula I can be formed as shown in Scheme I.The compound (i) can be halogenated with N-chlorosuccinamide,N-bromosuccinamide or N-iodosuccinamide to give compound (ii) whereX¹═Cl, Br, or I. The halo group of (ii) can be coupled to Cy-M, where Mis a boronic acid, boronic ester or an appropriately substituted metal(e.g., Cy-M is Cy-B(OH)₂, Cy-Sn(Bu)₄, or Zn-Cy), under standard Suzukiconditions or standard Stille conditions (e.g., in the presence of apalladium(0) catalyst, such as tetrakis(triphenylphosphine)palladium(0)and a base (e.g., a bicarbonate or carbonate base) or standard Negishiconditions (e.g., in the presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0), to give a derivative offormula (iii). Alternatively, Cy-M can be a cyclic amine (where M is Hand attached to the amine nitrogen) with coupling to compound (ii) beingperformed by heating in base or under Buchwald conditions (e.g., in thepresence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base (e.g., an alkoxidebase)) to afford ketone (iii). Reduction of the ketone (iii) with asuitable reagent, such as sodium tetrahydroborate can furnish thealcohol (iv) which can be converted to a derivative bearing a leavinggroup (v), (e.g., Lg is chloride via reaction with cyanuric chloride ormesylate via reaction with methanesulfonic anhydride). Finally, compound(v) can be reacted with 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine((vi)) under basic conditions (e.g., NaH or CsCO₃ or K₂CO₃) to give acompound of Formula I (vii).

Alternatively, compounds of Formula I can also be formed as shown inScheme II. The ketone compound (i) can be halogenated withN-chlorosuccinamide, N-bromosuccinamide or N-iodosuccinamide to givecompound (ii) where X¹═Cl, Br, or I. Ketone (ii) can be reduced with asuitable reagent, such as sodium tetrahydroborate, to give an alcohol(iii) which can be converted to a derivative bearing a leaving group,(e.g., Lg is chloride via reaction with cyanuric chloride or mesylatevia reaction with methanesulfonic anhydride) and then reacted with aheterocycle to give a heterocyclic derivative (iv). The enantiomers ofcompound (iv) can be separated by chiral chromatography to afford asingle enantiomer of heterocyclic compound (v). Finally, the halo groupof (v) can be coupled to Cy-M, where M is a boronic acid, boronic esteror an appropriately substituted metal (e.g., Cy-M is Cy-B(OH)2,Cy-Sn(Bu)4, or Zn-Cy), under standard Suzuki conditions or standardStille conditions (e.g., in the presence of a palladium(0) catalyst,such as tetrakis(triphenylphosphine)palladium(0) and a base (e.g., abicarbonate or carbonate base) or standard Negishi conditions (e.g., inthe presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0), to give a derivative ofFormula I (vi).

Compounds of Formula I can also be formed as shown in Scheme III. Thephenol (i) can be alkylated using Mitsunobu conditions (e.g., R′OH,DEAD, Ph₃P) or standard alkylating conditions (R′-Lg, Lg=leaving group)to afford ether derivatives (ii), respectively. The halo group of (ii)can be coupled to Cy-M, where M is a boronic acid, boronic ester or anappropriately substituted metal (e.g., Cy-M is Cy-B(OH)2, Cy-Sn(Bu)4, orZn-Cy), under standard Suzuki conditions or standard Stille conditions(e.g., in the presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base (e.g., a bicarbonateor carbonate base) or standard Negishi conditions (e.g., in the presenceof a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0), to give a derivative offormula (iii). Alternatively, Cy-M can be a cyclic amine (where M is Hand attached to the amine nitrogen) with coupling to compound (ii) beingperformed by heating in base or under Buchwald conditions (e.g., in thepresence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)-palladium(0) and a base (e.g., an alkoxidebase)) to afford compounds of formula (iii). The ketone (iii) can betransformed using similar methods as shown in Scheme I and II to affordcompounds of Formula I (iv). Alternatively, the halo-ketone (ii) can betransformed using similar methods as shown in Scheme I and II to affordhalo intermediate (v). Suzuki, Stille, Negishi or Buchwald coupling ofCy-M with halo intermediate (v) by similar methods described in SchemesI and II can also afford compounds of Formula I (vi).

Ketones which can be used in the processes of Scheme I, II and III, canbe formed as shown in Scheme IV below. The carboxylic acid (i) can beactivated with a coupling agent (e.g. HBTU or HATU) and then reactedwith N, O-dimethylhydroxylamine to give a N-methoxy-N-methylcarboxamide.The phenols can be alkylated using Mitsunobu conditions (e.g., R²OH,DEAD, Ph₃P) or standard alkylating conditions (R²-Lg, Lg=leaving group)to afford the ether derivatives (ii), respectively. The halo group of(ii) (X¹ is halo) can be coupled to Cy-M, where M is a boronic acid,boronic ester or an appropriately substituted metal (e.g., Cy-M isCy-B(OH)2, Cy-Sn(Bu)4, or Zn-Cy), under standard Suzuki conditions orstandard Stille conditions (e.g., in the presence of a palladium(0)catalyst, such as tetrakis(triphenylphosphine)palladium(O) and a base(e.g., a bicarbonate or carbonate base) or standard Negishi conditions(e.g., in the presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0), to give a derivative offormula (iii). Alternatively, Cy-M can be a cyclic amine (where M is Hand attached to the amine nitrogen) with coupling to compound (ii) beingperformed by heating in base or under Buchwald conditions (e.g., in thepresence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base (e.g., an alkoxidebase)) to afford amides (iii). Reaction of compound (iii) with aGrignard reagent of formula Me-MgX² (X²=halo) can give ketone (iv). Theketone (iv) can be transformed using similar methods as shown in SchemeI, II and III to afford compounds of Formula I.

Ketones which can be used in the processes of Scheme I, II and III, canalso be formed as shown in Scheme V below. The halo group (e.g., X¹═I)of (i) can be coupled to a zinc reagent Cy-Zn (e.g., such as tert-butyl3-iodoazetidine-1-carboxylate with Zn dust) under standardKnochel/Negishi conditions (e.g., in the presence of a palladium(0)catalyst, such as tri-(2-furyl)phosphine andtris(dibenzylideneacetone)dipalladium(0) and 1,2-dibromoethane andchlorotrimethylsilane) to give a derivative of formula (ii). Theazetidine (ii) can be deprotected (e.g., Pg=Boc, using TFA) and thenreacted under alkylating, acylating or reductive amination (e.g., R³Xsuch as R3-Br, R³COCl, R³—SO₂Cl, R³N═C═O or R³CHO and a reducing agent)conditions to afford ketone derivatives (iii) which can be converted tocompounds of Formula I (v) by similar methods shown in Schemes I, II,and III). Alternatively, the ketone (ii) can be reduced with suitablereagents (NaBH₄ or Corey's chiral CBS catalyst to give predominantly oneisomer of the alcohol), the resulting alcohol can be converted to aleaving group (e.g., Lg is chloride via reaction with cyanuric chlorideor mesylate via reaction with methanesulfonic anhydride) and then thechloride or mesylate reacted with an appropriate heterocycle (e.g.,similar to methods shown in Schemes I, II and III) to afford derivativesof formula (iv). The protecting group on the amine can be removed understandard conditions and then reacted under alkylating, acylating orreductive amination conditions (e.g., R³X such as R³—Br, R³COCl,R³—SO₂Cl, R³N═C═O or R³CHO and a reducing agent) to give compounds ofFormula I (v).

Compound of Formula I can be synthesized from an acid chloride compound(i) as illustrated in Scheme VI. Condensation of an acid chloride (i)with malononitrile in the presence of a base, such as sodium hydride,can give a dicyanoenol intermediate, which can be O-methylated with anappropriate reagent, such as dimethyl sulfate in the presence of anappropriate base, such as sodium bicarbonate, to yield an enol ether(ii). Reaction of enol ether (ii) with hydrazine dihydrochloride in thepresence of a suitable base, such as triethylamine, can give a pyrazolecompound (iii). Pyrazole compound (iii) can then be reacted withformamide to give pyrazolopyrimidine (iv). Finally, compound (iv) can bereacted with appropriate compound bearing a leaving group (v) underbasic conditions to give a compound of Formula I (vi).

Compounds of Formula I can also be formed as shown in Scheme VII. Thehalo group, X¹, of (i) can be coupled to an alkene (e.g., acrylate oracrylamide) under standard Heck conditions (e.g., in the presence of apalladium(II) catalyst, such as palladium acetate) to give an alkene offormula (ii). Reaction of alkene (ii) with nitromethane in the presenceof DBU can afford the nitro derivative (iii) which can be reduced understandard conditions (e.g., NiCl₂/NaBH₄) to give a free amine whichcyclizes to form lactam (iv). The lactam can be alkylated under standardconditions (e.g., R³—X², where X²=halo, in the presence of a base, suchas TEA or NaH) to give an N-alkyl-lactam (v). Compounds of formula (v),and pyrrolidines derived from the reduction of the lactam (v) withsuitable reducing agents, such as LiAlH₄, can be converted to compoundsof Formula I using conditions described in Schemes I, II and III.

Compounds of Formula I can also be formed as shown in Scheme VIII. Thehalo group X¹ of (i) can be coupled to an alkene boronic acid or esterunder standard Suzuki conditions (e.g., in the presence of apalladium(0) catalyst, such as tetrakis(triphenylphosphine)palladium(O))to give an alkene of formula (ii). Epoxidation of alkene (ii) with mCPBAcan afford the epoxide (iii) which can be reacted with a secondary orprimary amine (amine=NH₂R³) to give amino compounds of formula (iv).Secondary or tertiary amine derivatives (iv) can be further reacted withcarbonyldiamidazole or phosgene to form an oxazolidinone (v) or anacetyl-halide (e.g., chloro-acetylchloride in the presence of base, suchas TEA) to give the N-acyl derivative which can be converted to themorpholinone derivative (vi) upon treatment with a base (e.g., NaH).Compounds of formula (iv, v, and vi) can be deprotected using standardconditions (e.g., compounds protected with THP groups may be treatedwith an acid, such as TFA or HCl) to give compounds of Formula I.

Compounds of Formula I can also be formed as shown in Scheme IX.Sharpless amino-hydroxylation of an alkene of formula (i) under suitableconditions (A or B, as described in JACS, 2001, 123(9), 1862-1871 and JOrg. Chem, 2011, 76, 358-372) can give either amino-hydroxy isomer (ii)or (iii). Compounds (ii) and (iii) can be reacted withcarbonyldiamidazole or phosgene to form an oxazolidinone (iv), or anacetyl-halide (e.g., chloro-acetylchloride in the presence of base, suchas TEA) to give an N-acyl derivative which can be converted to themorpholinone derivative (v) upon treatment with a base (e.g., NaH). Thealternate amino-hydroxy isomer (iii) can be converted to oxazolidinoneand morpholinone derivatives as shown in Scheme XV.

Compounds of Formula I can be synthesized as shown in Scheme X. The halogroup (e.g., X¹═Cl, Br, I) of (i) can be converted to the boronate ester(ii) under standard conditions (e.g., pinnacle boronate ester in thepresence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0)). Boronate (ii) can be reactedwith an arylhalide or heteroarylhalide (e.g., R³—X²) under Suzukiconditions (e.g., in the presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base, such as Na₂CO₃) togive formula (iii). Formula (iii) can be converted to Formula I usingthe reaction conditions described in Schemes I, II or III.

Compounds of Formula I, where R⁴═F or CN, can be formed as shown inScheme XI. Compound (i) can be acylated with a suitable acylatingreagent (e.g., Me-COCl) to form an ester which can be rearranged underLewis acid conditions (e.g., BF₃/HOAc complex) to afford ketone (ii).Ketone (ii) can be halogenated with N-chlorosuccinamide,N-bromosuccinamide or N-iodosuccinamide to give phenol (iii), whereX¹═Cl, Br, or I. Compound (iii) can be alkylated (e.g. R²—X and a base,such as NaH or Na₂CO₃; or under Mitsunobu conditions) to afford theether (iv). The fluoro group of (iv) can be displaced (e.g., with NaCNor KCN) to give cyano derivative (v). The halo group of (v) can becoupled to Cy-M, where M is a boronic acid, boronic ester or anappropriately substituted metal (e.g., Cy-M is Cy-B(OH)2, Cy-Sn(Bu)4, orZn-Cy), under standard Suzuki conditions or standard Stille conditions(e.g., in the presence of a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base (e.g., a bicarbonateor carbonate base) or standard Negishi conditions (e.g., in the presenceof a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0)), to give a derivative offormula (vi). Alternatively, Cy-M can be a cyclic amine (where M is Hand attached to the amine nitrogen) and coupled to compound (v) byheating in base or under Buchwald conditions (e.g., in the presence of apalladium(0) catalyst, such as tetrakis(triphenylphosphine)palladium(0)and a base (e.g., an alkoxide base)) to afford ketone (vi). Reduction ofthe ketone (vi) with a suitable reagent, such as sodium tetrahydroborateor the Corey CBS reagent can furnish the alcohol which can be convertedto a derivative bearing a leaving group, (e.g., Lg is chloride viareaction with cyanuric chloride or mesylate via reaction withmethanesulfonic anhydride) and then reacted with3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine under basic conditions(e.g., NaH or CsCO₃ or K₂CO₃) to give a compound of Formula I (viii).Alternatively, the last two steps can be inverted so that the ketone (v)can be reduced to give an alcohol which is converted to a leaving groupand displaced with the heterocycle first and then the Suzuki, Stille,Negishi or Buchwald coupling is performed to give compounds of Formula I(viii). The fluoro derivatives (iv) can also be converted to compoundsof Formula I by eliminating the cyanation step in Scheme XI.

Compounds of Formula I can also be formed as shown in Scheme XII.Compound (i) can be acylated with a suitable acylating reagent (e.g.,Me-COCl) to form an ester which can be rearranged under Lewis acidconditions (e.g., AlCl₃ or BF₃/HOAc complex) to afford ketone (ii).Halogenation of ketone (ii) using NX¹S (e.g., NX¹S═N-chlorosuccinamide,N-bromosuccinamide or N-iodosuccinamide) can give compound (iii), whereX¹═Cl, Br, or I. The phenol can be converted to an ether (iv) usingstandard conditions (e.g., inorganic base, such as K₂CO₃, and an alkylhalide, such as Et-I). The halo group of (iv) can be coupled to R³-M,where M is a boronic acid, boronic ester or an appropriately substitutedmetal (e.g., R³-M is R³—B(OH)₂, R³—Sn(Bu)₄, or Zn—R³ and R³ is asubstituted or unsubstituted olefin, such as vinyl) under standardSuzuki conditions or standard Stille conditions (e.g., in the presenceof a palladium(0) catalyst, such astetrakis(triphenylphosphine)palladium(0) and a base (e.g., a bicarbonateor carbonate base) to give a derivative of formula (v). The alkene canthen be dihydroxylated using Sharpless conditions to afford the diol(vi). Enhancement of one enantiomer of the secondary alcohol can beachieved using standard Sharpless asymmetric dihydroxylation methods.The secondary alcohol can be converted to the N-Boc protected amine viaa 6 step process (e.g. silyl protection (e.g., TBS-Cl and DIEA) of theprimary alcohol, mesylation of the secondary alcohol, displacement ofthe mesylate with NaN₃, reduction of the azide with Ph₃P, Boc protectionof the resulting primary amine and then deprotection of the silylprotecting group on the primary alcohol with TBAF) to affordamino-alcohol (vii). The amino-alcohol (vii) can be converted into theoxazolidinone by treatment with phosgene and subsequent reduction of theketone with a suitable reagent, such as sodium tetrahydroborate orsodium borohydride can furnish the alcohol (viii) which can be convertedto a derivative bearing a leaving group (ix) (e.g., Lg is chloride viareaction with cyanuric chloride or mesylate via reaction withmethanesulfonic anhydride). Finally, compound (ix) can be reacted withan appropriate heterocycle (x) (e.g.,3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine or4-aminopyrido[2,3-d]pyrimidin-5(8H)-one) under basic conditions (e.g.,NaH or Cs₂CO₃ or K₂CO₃) to give a compound of Formula I (xi).

The invention will be described in greater detail by way of specificexamples. The following examples are offered for illustrative purposes,and are not intended to limit the invention in any manner. Those ofskill in the art will readily recognize a variety of non-criticalparameters which can be changed or modified to yield essentially thesame results. The compounds of the Examples have been found to be PI3Kinhibitors according to at least one assay described herein.

EXAMPLES

The example compounds below containing one or more chiral centers wereobtained in racemate form or as isomeric mixtures, unless otherwisespecified. Salt stoichiometry which is indicated any of the productsbelow is meant only to indicate a probable stoichiometry, and should notbe construed to exclude the possible formation of salts in otherstoichiometries. The abbreviations “h” and “min” refer to hour(s) andminute(s), respectively.

Example 1.1-{1-[5-Chloro-3-(1-isopropylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

Step 1. 1-(5-Chloro-2-hydroxy-3-iodo-4-methylphenyl)ethanone

To a stirred solution of 1-(5-chloro-2-hydroxy-4-methylphenyl)ethanone(from Oakwood, 50.0 g, 271 mmol) in acetic acid (300 mL) was addedN-iodosuccinimide (73.1 g, 325 mmol) and the resulting mixture wasstirred on a heating mantle between 60-80° C. over 3.5 hours then cooledto room temperature and stirred overnight. Water (500 mL) was added tothe mixture in portions, which caused a dark solid to form. Afterstirring for 10 minutes, the solids were filtered, washing withadditional water. The light to dark brown solids were dried under vacuumfor 4 hours then air dried over the weekend to give 81.3 g (97%) of thedesired product. LCMS calculated for C₉H₉ClIO₂ (M+H)⁺: m/z=310.9; Found:311.0. ¹H NMR (300 MHz, CDCl₃): δ 13.21 (s, 1H), 7.71 (s, 1H), 2.65 (s,3H), 2.63 (s, 3H) ppm.

Step 2. 1-(5-Chloro-3-iodo-2-methoxy-4-methylphenyl)ethanone

Potassium carbonate (72.4 g, 524 mmol) was added to a mixture of1-(5-chloro-2-hydroxy-3-iodo-4-methylphenyl)ethanone (81.3 g, 262 mmol)and methyl iodide (19.6 mL, 314 mmol) in N,N-dimethylformamide (250 mL).The mixture was stirred at room temperature for 4 hours. Water (500 mL)was added and stirred for 15 minutes. The dark solids were filtered anddried in vacuo to give 42.3 g of the desired product. The filtrate wasextracted with EtOAc (4×). The combined filtrates were washed with water(2×) and brine, dried (MgSO₄), filtered and concentrated. The solidswere dried in vacuo to give an additional 37.2 g of the desired product.The product was used without further purification. LCMS calculated forC₁₀H₁₁ClIO₂ (M+H)⁺: m/z=324.9; Found: 325.0. ¹H NMR (300 MHz, CDCl₃): δ7.62 (s, 1H), 3.78 (s, 3H), 2.65 (s, 3H), 2.62 (s, 3H) ppm.

Step 3. tert-Butyl3-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)azetidine-1-carboxylate

Zinc (1.71 g, 26.2 mmol) was suspended in N,N-dimethylformamide (45.0mL) and 1,2-dibromoethane (210 μL, 2.5 mmol) was added. The mixture washeated at 60° C. for 10 minutes and then cooled to room temperature.Chlorotrimethylsilane (330 μL, 2.6 mmol) was added and stirred at 60° C.for 10 minutes and cooled to room temperature. A solution of tert-butyl3-iodoazetidine-1-carboxylate (from Oakwood, 6.25 g, 22.1 mmol) inN,N-dimethylformamide (5.0 mL) was then added and the mixture stirred atroom temperature for 1 hour.1-(5-chloro-3-iodo-2-methoxy-4-methylphenyl)ethanone (5.00 g, 15.4mmol), tri-(2-furyl)phosphine (358 mg, 1.54 mmol), andtris(dibenzylideneacetone)dipalladium(0) (0.70 g, 0.77 mmol) were addedin order and the reaction mixture was warmed to 70° C. and stirredovernight. The mixture was cooled to room temperature and partitionedbetween ethyl acetate (EtOAc) and sat. NH₄Cl solution. The layers wereseparated and the aqueous extracted further with EtOAc (2×). Thecombined organics were washed with water and brine, dried over MgSO₄,and concentrated. The residue was purified on silica gel, eluting with0-30% EtOAc in hexanes to give 3.0 g (55%) of the desired product as anorange solid. LCMS calculated for C₁₈H₂₄ClNO₄Na (M+Na)⁺: m/z=376.1;Found: 376.0. ¹H NMR (400 MHz, CDCl₃): δ 7.52 (s, 1H), 4.32, (m, 2H),4.16 (m, 3H), 3.66 (s, 3H), 2.59 (s, 3H), 2.31 (s, 3H), 1.45 (s, 9H)ppm.

Step 4. tert-Butyl3-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]azetidine-1-carboxylate

To a solution of tert-butyl3-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)azetidine-1-carboxylate(1.3 g, 3.7 mmol) in methanol (20 mL) stirring at 0° C. was added sodiumtetrahydroborate (0.167 g, 4.41 mmol). The mixture was stirred at 0-5°C. for 1 hour. The reaction was quenched with water and extracted withEtOAc (3×). The combined extracts were dried over MgSO₄, filtered andconcentrated to give 1.3 g (100%) of the desired product. LCMScalculated for C₁₈H₂₆ClNO₄Na (M+Na)⁺: m/z=378.2; Found: 378.1. ¹H NMR(400 MHz, CDCl₃): δ 7.37 (s, 1H), 5.10 (q, 1H), 4.30 (m, 2H), 4.14 (m,3H), 3.63 (s, 3H), 2.25 (s, 3H), 1.48 (d, 3H), 1.44 (s, 9H) ppm.

Step 5. tert-Butyl3-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]azetidine-1-carboxylate

Cyanuric chloride (from Aldrich, 1.22 g, 6.62 mmol) was weighed into aflask and N,N-dimethylformamide (0.512 mL, 6.62 mmol) was added. Afterstirring for a few minutes a solution of tert-butyl3-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]azetidine-1-carboxylate(1.5 g, 4.2 mmol) in methylene chloride (30 mL) was added. The resultingmixture was stirred at room temperature overnight. Water was added, andthen diluted with dichloromethane. The layers were separated and theorganics were washed with sat. NaHCO₃ solution, water, brine, dried overMgSO₄, and concentrated. The resulting residue was purified on silicagel, eluting with 0-35% EtOAc in hexanes to give the desired product(1.36 g, 86%). LCMS calculated for C₁₃H₁₇ClNO (M-Cl-Boc+H)⁺: m/z=238.1;Found: 238.1. ¹H NMR (400 MHz, CDCl₃): δ 7.46 (s, 1H), 5.44, (q, 1H),4.32 (m, 2H), 4.18-4.10 (m, 3H), 3.67 (s, 3H), 2.27 (s, 3H), 1.79 (d,3H), 1.44 (s, 9H) ppm.

Step 6. tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidine-1-carboxylate

At room temperature, sodium hydride (0.32 g, 8.0 mmol) was added to asuspension of 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (fromChemBridge, 0.59 g, 4.0 mmol) in N,N-dimethylformamide (20 mL). Theresulting mixture was stirred at room temperature for 25 minutes duringwhich time the suspension became a nearly clear solution. To theresultant mixture was added a solution of tert-butyl3-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]azetidine-1-carboxylate(1.35 g, 3.61 mmol, from Example 1, step 5) in N,N-dimethylformamide (10mL). The mixture was stirred at 50° C. overnight. After cooling, themixture was diluted with water and extracted with EtOAc (2×). Thecombined extracts were washed with water and brine, dried over MgSO₄ andconcentrated. The resulting residue was purified on silica gel, elutedwith 0-10% MeOH in dichloromethane to give 1.03 g (59%) of the desiredproduct as a yellow gum. The racemic products were applied on aPhenomenex Lux-Cellulose 2 column (21.1×250 mm, 5 micron particle size),eluting with 10% ethanol in hexanes at a flow rate of 18 mL/min, 4mg/injection, to provide two enantiomers. The retention time of thefirst peak was 8.34 min and the retention time for the second peak was10.92 min. Peak 1 (463 mg), LCMS calculated for C₂₄H₃₂ClN₆O₃(M+H)⁺:m/z=487.2; Found: 487.1. ¹H NMR (400 MHz, CDCl₃): δ 8.21 (s, 1H), 7.37(s, 1H), 6.30, (q, 1H), 5.40 (s, 2H), 4.23 (m, 2H), 4.17-4.00 (m, 3H),3.57 (s, 3H), 2.58 (s, 3H), 2.16 (s, 3H), 1.76 (d, 3H), 1.37 (s, 9H)ppm.

Step 7.1-[1-(3-Azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

To a solution of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidine-1-carboxylate(318 mg, 0.653 mmol) (peak 1 from above) in methylene chloride (3.2 mL)was added 4.0 M hydrogen chloride in 1,4-dioxane (1.6 mL, 6.5 mmol). Theresulting mixture was stirred at room temperature for 75 minutes. Thesolvents were evaporated and the residue dried in vacuo to give 0.30 gof the desired product as the bis-HCl salt. LCMS calculated forC₁₉H₂₄ClN₆O (M+H)⁺: m/z=387.2; Found: 387.1.

Step 8.1-{-[5-Chloro-3-(1-isopropylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (58 mg, 0.13 mmol), acetone (18.5 μL, 0.252 mmol) andtriethylamine (54.5 μL, 0.391 mmol) in methylene chloride (1.0 mL) wasadded resin of sodium triacetoxyborohydride (108 mg, 0.249 mmol). Theresulting mixture was stirred for 3 hours at room temperature. Themixture was filtered and concentrated. The crude product was purifiedusing RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive 50 mg (60%) of the desired product as the TFA salt. LCMS calculatedfor C₂₂H₃₀ClN₆O (M+H)⁺: m/z=429.2; Found: 429.1. The product wasisolated as a single enantiomer. ¹H NMR (500 MHz, DMSO-d₆): δ 8.47 (s,1H), 7.46 (s, 1H), 6.29 (q, J=6.9 Hz, 1H), 4.52 (m, 2H), 4.21 (m, 1H),4.15 (t, J=9.8 Hz, 1H), 4.06 (t, J=9.7 Hz, 1H), 3.53 (s, 3H), 3.39-3.27(m, 1H), 2.61 (s, 3H), 2.11 (s, 3H), 1.75 (d, J=6.8 Hz, 3H), 1.11 (dd,J=6.0, 3.8 Hz, 6H) ppm.

Example 2.1-{1-[3-(1-Acetylazetidin-3-yl)-5-chloro-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminetrifluoroacetate

Step 1.1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

To a solution of the racemic tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidine-1-carboxylate(146 mg, 0.300 mmol) (racemic intermediate from Example 1 Step 6) inmethylene chloride (1.5 mL) was added 4.0 M hydrogen chloride in1,4-dioxane (0.75 mL, 3.0 mmol). After stirred at rt for 2 h, thesolvents were evaporated and the resulting residue dried in vacuo togive 138 mg of the desired product as the HCl salt. LCMS calculated forC₁₉H₂₄ClN₆O (M+H)⁺: m/z=387.2; Found: 387.1.

Step 2.1-{1-[3-(1-Acetylazetidin-3-yl)-5-chloro-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminetrifluoroacetate

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (20.0 mg, 0.0435 mmol, from Example 2, step 1) andtriethylamine (30.3 μL, 0.217 mmol) in methylene chloride (0.20 mL) wasadded acetyl chloride (6.18 μL, 0.0870 mmol). The resulting mixture wasstirred overnight at room temperature. The solvents were evaporated andthe crude purified using RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.05% TFA, at flow rate of 30mL/min) to give the desired product as the TFA salt. The product wasisolated as a racemic mixture. LCMS calculated for C₂₁H₂₆ClN₆O₂(M+H)⁺:m/z=429.2; Found: 429.1. ¹H NMR (400 MHz, DMSO-d₆): δ 8.35 (s, 1H), 7.34(s, 1H), 6.26 (q, 1H), 4.50 (m, 1H), 4.28-4.20 (m, 2H), 4.01 (m, 1H),3.88 (m, 1H), 3.52 (s, 3H), 2.58 (s, 3H), 2.18 (s, 3H), 1.75-1.71 (m,6H) ppm.

Example 3.1-{1-[5-Chloro-2-methoxy-4-methyl-3-(1-propionylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminetrifluoroacetate

This compound was prepared using procedures analogous to those forExample 2, with propanoyl chloride instead of acetyl chloride. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₂H₂₈ClN₆O₂(M+H)⁺: m/z=443.2; Found: 443.2. ¹H NMR (400 MHz, DMSO-d₆) δ8.30 (s, 1H), 7.33 (s, 1H), 6.25 (q, 1H), 4.49 (m, 1H), 4.27-4.18 (m,2H), 4.02 (m, 1H), 3.90 (m, 1H), 3.54 (s, 3H), 2.57 (s, 3H), 2.18 (s,3H), 2.05 (q, 2H), 1.72 (d, 3H), 0.93 (t, 3H) ppm.

Example 4.1-(1-{5-Chloro-3-[1-(cyclopropylmethyl)azetidin-3-yl]-2-methoxy-4-methylphenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and cyclopropanecarboxaldehyde(from Aldrich) instead of acetone. The product was isolated as a racemicmixture. LCMS calculated for C₂₃H₃₀ClN₆O (M+H)⁺: m/z=441.2; Found:441.1. ¹H NMR (400 MHz, DMSO-d₆): δ 8.06 (s, 1H), 7.13 (s, 1H), 5.96 (q,1H), 4.22 (m, 2H), 4.07 (m, 1H), 3.90 (m, 1H), 3.80 (m, 1H), 3.24 (s,3H), 2.68 (t, 2H), 2.21 (s, 3H), 1.80 (s, 3H), 1.45 (d, 3H), 0.64 (m,1H), 0.24 (m, 2H), 0.01 (m, 2H) ppm.

Example 5.1-{1-[5-chloro-2-methoxy-4-methyl-3-(1-methylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and formaldehyde instead ofacetone. The crude purified using RP-HPLC (XBridge C18 column, elutingwith a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₀H₂₆ClN₆O (M+H)⁺: m/z=401.2; Found: 401.2.

Example 6.1-{1-[5-Chloro-3-(1-ethylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and acetaldehyde instead ofacetone. The crude purified using RP-HPLC (XBridge C18 column, elutingwith a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₁H₂₈ClN₆O (M+H)⁺: m/z=415.2; Found: 415.1.

Example 7.1-{1-[5-Chloro-3-(1-isobutylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and isobutyraldehyde instead ofacetone. The crude purified using RP-HPLC (XBridge C18 column, elutingwith a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₃H₃₂ClN₆O (M+H)⁺: m/z=443.2; Found: 443.1. ¹H NMR (400 MHz, CDCl₃): δ8.29 (s, 1H), 7.38 (s, 1H), 6.37 (q, 1H), 5.37 (s, 2H), 4.01 (m, 2H),3.87 (m, 1H), 3.57 (s, 3H), 3.05 (t, 1H), 2.86 (t, 1H), 2.64 (s, 3H),2.18 (d, 2H), 2.11 (s, 3H), 1.82 (d, 3H), 1.62 (m, 1H), 0.89 (d, 6H)ppm.

Example 8.1-{1-[3-(1-sec-butylazetidin-3-yl)-5-chloro-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and 2-butanone instead ofacetone. The crude was purified using RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a mixture of diastereomers. LCMS calculated forC₂₃H₃₂ClN₆O (M+H)⁺: m/z=443.2; Found: 443.1.

Example 9.1-(1-{5-Chloro-2-methoxy-3-[1-(2-methoxyethyl)azetidin-3-yl]-4-methylphenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared using procedures analogous to those forExample 1, with racemic1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride from Example 2, Step 1 and methoxyacetaldehyde insteadof acetone. The crude was purified using RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₂H₃₀ClN₆O₂(M+H)⁺: m/z=445.2; Found: 445.2.

Example 10.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-methylazetidine-1-carboxamide

This compound was prepared using procedures analogous to those forExample 2, with methyl isocyanate instead of acetyl chloride The crudepurified using RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product. The product was isolated as aracemic mixture. LCMS calculated for C₂₁H₂₇ClN₇O₂(M+H)⁺: m/z=444.2;Found: 444.2.

Example 11.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

Step 1. 1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone

To a stirred solution of 1-(5-chloro-2-methoxy-4-methylphenyl)ethanone(5.00 g, 25.2 mmol, from Oakwood) in acetic acid (100 mL) was addedN-bromosuccinimide (4.93 g, 27.7 mmol) and the resulting mixture heatedat 100° C. for 18 hours. After cooling to ambient temperature, thereaction mixture was concentrated in vacuo, then neutralized with sat.sodium bicarbonate, filtered off insoluble succinimide. The filtrate wasextracted with EtOAc. The combined organic layers were washed withbrine, dried over sodium sulfate, and then concentrated to dryness underreduced pressure. The residue was purified on silica gel, eluting with 0to 50% EtOAc in hexanes, to give the desired products (2.66 g, 38%).LCMS calculated for C₁₀H₁₁BrClO₂ (M+H)⁺: m/z=277.0; found: 277.0. ¹H NMR(DMSO-d₆, 300 MHz): δ 7.70 (1H, s), 3.77 (3H, s), 2.57 (3H, s), 2.50(3H, s) ppm.

Step 2.5-(3-Acetyl-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpyridine-2-carboxamide

To a mixture of 1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone(0.38 g, 1.4 mmol) andN,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carboxamide(from PepTech, 0.46 g, 1.6 mmol) in 1,4-dioxane (6 mL), potassiumcarbonate (0.38 g, 2.7 mmol) in water (2 mL) was added. The reactionmixture was bubbled with N₂. Tetrakis(triphenylphosphine)palladium(0)(0.095 g, 0.082 mmol) was added and the reaction was stirred overnightat 100° C. The reaction was diluted with water, extracted with EtOAc.The combined organic layers were dried over MgSO₄, concentrated andpurified on silica gel (eluting with 0-100% EtOAc in hexanes) to givethe desired product. LCMS calculated for C₁₈H₂₀ClN₂O₃ (M+H)⁺: m/z=347.1;Found: 347.1.

Step 3.5-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide

To a solution of5-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpyridine-2-carboxamide(106 mg, 0.306 mmol) in methanol (2 mL) cooled at 0° C. was added sodiumtetrahydroborate (14 mg, 0.37 mmol). The mixture was stirred at roomtemperature for 1 hour, then quenched with water, extracted with EtOAc.The organic layers were dried over MgSO₄ and concentrated to give crudealcohol. LCMS calculated for C₁₈H₂₂ClN₂O₃(M+H)⁺: m/z=349.1; Found:349.1.

Step 4.5-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide

Cyanuric chloride (85 mg, 0.46 mmol) was added to N,N-dimethylformamide(0.036 mL, 0.46 mmol) at room temperature. After the formation of awhite solid (10 minutes), methylene chloride (2 mL) was added, followedby5-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(115 mg, 0.330 mmol, from Example 11, step 3). After the addition, themixture was stirred at room temperature overnight. Water was added, andthen diluted with dichloromethane. The organic phase was washed withsat. NaHCO₃ solution, water and brine, then dried over MgSO₄,concentrated. The residue was purified on silica gel (eluting with 0 to80% EtOAc in hexanes) to give the desired product (76 mg, 63%). LCMScalculated for C₁₈H₂₁Cl₂N₂O₂ (M+H)⁺: m/z=367.1; Found: 367.0.

Step 5.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

To a solution of 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (6.1 mg,0.041 mmol) in N,N-dimethylformamide (0.4 mL) was added sodium hydride(60%, 2.0 mg, 0.082 mmol) at 0° C. and the mixture was stirred at roomtemperature for 10 minutes. To the resultant mixture was added asolution of5-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(15.0 mg, 0.0408 mmol) in N,N-dimethylformamide (0.2 mL). The mixturewas stirred at room temperature overnight. The crude mixture waspurified on RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as bis-TFA salt. The product was isolated as aracemic mixture. LCMS calculated for C₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2;Found: 480.1.

Example 13.1-{1-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

Step 1. 1-(5-Chloro-4-fluoro-2-hydroxyphenyl)ethanone

To 4-chloro-3-fluorophenol (from Aldrich, 20 g, 100 mmol) was addedacetyl chloride (14.1 mL, 199 mmol) under N₂ with stirring. Theresulting mixture turned into a clear solution at room temperaturequickly and it was heated at 60° C. for 2 hours. To the resultantmixture was added aluminum trichloride (25.0 g, 187 mmol) in portionsand the reaction mixture was heated at 180° C. for 30 minutes. Thesolids slowly dissolved at high temperature. The reaction mixture wasthen cooled to room temperature while the flask was swirled carefully inorder for the solid to form a thin layer inside the flask and thenslowly quenched with 1.0 N HCl (300 mL) while cooling in an ice-bath andstirred overnight. The yellow precipitate was washed with water anddried under vacuum to give the desired product as a yellow solid (23.8g), which was directly used in the next step without furtherpurification.

Step 2. 1-(5-Chloro-4-fluoro-2-hydroxy-3-iodophenyl)ethanone

A solution of 1-(5-chloro-4-fluoro-2-hydroxyphenyl)ethanone (23.8 g, 126mmol) in acetic acid (100 mL) was treated with N-iodosuccinimide (34.1g, 151 mmol) and stirred at 70° C. for 2 hr. The reaction mixture wasconcentrated, diluted with EtOAc and quenched with sat. NaHCO₃ solutionuntil the bubbling stopped. The organic layers were separated, washedwith water, dried over MgSO₄ and stripped to give the desired productwhich was used in the next step without further purification.

Step 3. 1-(5-Chloro-4-fluoro-3-iodo-2-methoxyphenyl)ethanone

1-(5-Chloro-4-fluoro-2-hydroxy-3-iodophenyl)ethanone (13 g, 41 mmol) wasdissolved in N,N-dimethylformamide (41.3 mL). Methyl iodide (3.9 mL, 62mmol) was added followed by potassium carbonate (11 g, 83 mmol). Thereaction was heated at 60° C. for 1 hour. The mixture was cooled to roomtemperature, diluted with ether. The organic layers were separated andcombined, washed with water, dried over MgSO₄, concentrated and purifiedon silica gel (eluting with 0 to 10% EtOAc in hexanes) to give thedesired product (10 g, 70%). LCMS calculated for C₉HsClFIO₂ (M+H)⁺:m/z=328.9; Found: 328.9.

Step 4. tert-Butyl3-(3-acetyl-5-chloro-6-fluoro-2-methoxyphenyl)azetidine-1-carboxylate

Zinc (0.682 g, 10.4 mmol) was suspended with 1,2-dibromoethane (0.0598mL, 0.694 mmol) in N,N-dimethylformamide (12 mL). The mixture was heatedat 70° C. for 10 minutes and then cooled to room temperature.Chlorotrimethylsilane (0.088 mL, 0.69 mmol) was added dropwise andstirring was continued for 1 hour. A solution of tert-butyl3-iodoazetidine-1-carboxylate (2.5 g, 8.7 mmol) in N,N-dimethylformamide(10 mL) was then added and the mixture was heated at 40° C. for 1 hourbefore a mixture of 1-(5-chloro-4-fluoro-3-iodo-2-methoxyphenyl)ethanone(3.0 g, 9.1 mmol), tris(dibenzylideneacetone)dipalladium(0) (0.16 g,0.17 mmol) and tri-(2-furyl)phosphine (0.081 g, 0.35 mmol) inN,N-dimethylformamide (20 mL) was added. The reaction mixture was warmedto 70° C. and stirred overnight. The mixture was then cooled to roomtemperature and partitioned between ether and sat. NH₄Cl solution. Theorganic layers were washed with water, dried over MgSO₄, concentratedand purified on silica gel (eluting with 0 to 25% EtOAc in hexanes) togive the desired product (0.8 g). LCMS calculated for C₁₇H₂₁ClFNO₄Na(M+Na)⁺: m/z=380.1; Found: 380.1.

Step 5. tert-Butyl3-[3-chloro-2-fluoro-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate

To a solution of tert-butyl3-(3-acetyl-5-chloro-6-fluoro-2-methoxyphenyl)azetidine-1-carboxylate(0.17 g, 0.48 mmol) in methanol (3 mL) cooled at 0° C. was added sodiumtetrahydroborate (0.022 g, 0.57 mmol). The mixture was stirred at roomtemperature for 1 hour, then quenched with water, extracted with EtOAc.The organic layers were combined, dried over MgSO₄ and concentrated togive the crude alcohol (0.19 g). LCMS calculated for C₁₇H₂₃ClFNO₄Na(M+Na)⁺: m/z=382.1; Found: 382.0.

Step 6. tert-Butyl3-[3-chloro-5-(1-chloroethyl)-2-fluoro-6-methoxyphenyl]azetidine-1-carboxylate

Cyanuric chloride (140 mg, 0.78 mmol) was added to N,N-dimethylformamide(0.059 mL, 0.77 mmol) at room temperature. After the formation of awhite solid (ca. 10 minutes), methylene chloride (4 mL) was added,followed by tert-butyl3-[3-chloro-2-fluoro-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate(197 mg, 0.547 mmol). After addition, the mixture was stirred at roomtemperature overnight. Water was added, and then diluted withdichloromethane. The organic phases were washed with sat. NaHCO₃solution, water and brine, dried over MgSO₄, and concentrated. Theresulting residue was purified on silica gel (eluting with 0 to 30%EtOAc in hexanes) to give the desired product (110 mg, 53%).

Step 7. tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidine-1-carboxylate

To a solution of 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (7.9 mg,0.053 mmol) in N,N-dimethylformamide (0.6 mL) was added sodium hydride(60%, 2.5 mg, 0.11 mmol) at 0° C. and the mixture was stirred at roomtemperature for 10 minutes. To the mixture was added a solution oftert-butyl3-[3-chloro-5-(1-chloroethyl)-2-fluoro-6-methoxyphenyl]azetidine-1-carboxylate(20 mg, 0.053 mmol) in N,N-dimethylformamide (0.3 mL). The reactionmixture was stirred at 35° C. overnight, then quenched with water,extracted with ether. The combined organic layers were dried over MgSO₄and concentrated to afford the desired product which was used in nextstep directly. LCMS calculated for C₂₃H₂₉ClFN₆O₃ (M+H)⁺: m/z=491.2;Found: 491.1.

Step 8.1-{1-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

A mixture of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidine-1-carboxylate(14 mg, 0.028 mmol) in methylene chloride (0.2 mL) was treated with 4.0M hydrogen chloride in dioxane (0.2 mL, 0.8 mmol) at room temperaturefor 1 hour and then the solvent removed to give1-[1-(3-azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amineHCl salt. To a mixture of the crude HCl salt in acetonitrile (0.1mL)/methanol (0.1 mL)/tetrahydrofuran (0.1 mL) was addedN,N-diisopropylethylamine (0.1 mL, 0.6 mmol), followed by acetone (0.050mL, 0.68 mmol). The mixture was stirred for 30 minutes before theaddition of sodium triacetoxyborohydride (0.030 g, 0.14 mmol). Thereaction was stirred at room temperature overnight, then quenched andpurified on RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as TFA salt. The product was isolated as aracemic mixture. LCMS calculated for C₂₁H₂₇ClFN₆O (M+H)⁺: m/z=433.2;Found: 433.1.

Example 14.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

Step 1. 1-(5-Chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanone

1-(5-Chloro-2-hydroxy-3-iodo-4-methylphenyl)ethanone (18.9 g, 60.9 mmol)(from Example 1, Step 1) was dissolved in N,N-dimethylformamide (60.8mL). Iodoethane (7.3 mL, 91 mmol) was added followed by potassiumcarbonate (17 g, 120 mmol). The reaction was heated at 60° C. for 1hour. The mixture was cooled to room temperature, diluted with ether.The organic layers were combined, washed with water, dried over MgSO₄,concentrated and purified on silica gel (eluting with 0-10% EtOAc inhexanes) to give the desired product (18.9 g, 91.7%). LCMS calculatedfor Cl₁H₁₃ClIO₂ (M+H)⁺: m/z=339.0; Found: 339.0.

Step 2.5-(3-Acetyl-5-chloro-2-ethoxy-6-methylphenyl)-N,N-dimethylpyridine-2-carboxamide

To a mixture of 1-(5-chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanone(0.69 g, 2.0 mmol) andN,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carboxamide(0.68 g, 2.4 mmol) in 1,4-dioxane (10 mL), potassium carbonate (0.56 g,4.1 mmol) in water (3 mL, 200 mmol) was added. The reaction was bubbledwith N₂. Tetrakis(triphenylphosphine)palladium(0) (0.24 g, 0.20 mmol)was added and N₂ was bubbled. Reaction was stirred overnight at 95° C.The reaction was diluted with water, extracted with EtOAc. The combinedorganic layers were dried over MgSO₄, concentrated and purified onsilica gel (eluting with 0 to 90% EtOAc in hexanes) to give the desiredproduct (0.6 g, 82%). LCMS calculated for C₁₉H₂₂ClN₂O₃(M+H)⁺: m/z=361.1;Found: 361.0.

Step 3.5-[3-Chloro-6-ethoxy-5-(1-hydroxyethyl)-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide

To a solution of5-(3-acetyl-5-chloro-2-ethoxy-6-methylphenyl)-N,N-dimethylpyridine-2-carboxamide(0.60 g, 1.7 mmol) in methanol (10 mL) cooled at 0° C. was added sodiumtetrahydroborate (0.075 g, 2.0 mmol). The mixture was stirred at roomtemperature for 1 hour, then quenched with water, extracted with EtOAc.The extracts were dried over MgSO₄ and concentrated to give crudealcohol (0.6 g). LCMS calculated for C₁₉H₂₄ClN₂O₃(M+H)⁺: m/z=363.1;Found: 363.0.

Step 4.5-[3-Chloro-5-(1-chloroethyl)-6-ethoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide

Cyanuric chloride (0.43 g, 2.3 mmol) was added to N,N-dimethylformamide(0.18 mL, 2.3 mmol) at room temperature. After the formation of a whitesolid (10 minutes), methylene chloride (10 mL) was added, followed by5-[3-chloro-6-ethoxy-5-(1-hydroxyethyl)-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(0.6 g, 2 mmol). After addition, the mixture was stirred at roomtemperature overnight, then diluted with dichloromethane and washed withsat. NaHCO₃ solution. The organic layers were dried over MgSO₄,concentrated. The residue was purified on silica gel (eluting with 0 to50% EtOAc in hexanes) to give the desired product (0.58, 90%). LCMScalculated for C₁₉H₂₃Cl₂NO₂ (M+H)⁺: m/z=381.1; Found: 381.0.

Step 5.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

To a solution of 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (47 mg,0.31 mmol) in N,N-dimethylformamide (3 mL) was added sodium hydride(60%, 12.6 mg, 0.524 mmol) at 0° C. and the resultant mixture wasstirred at room temperature for 10 minutes. To the mixture was added asolution of5-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(100 mg, 0.3 mmol, from Example 14, step 4) in N,N-dimethylformamide (1mL). The reaction was stirred at 35° C. overnight. The reaction wasquenched and applied on RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.05% TFA, at flow rate of 30mL/min) to give the desired product as bis-TFA salt. The product wasisolated as a racemic mixture. LCMS calculated for C₂₅H₂₉ClN₇O₂(M+H)⁺:m/z=494.2; Found: 494.1.

Example 16.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide

Step 1. 1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl) ethanol

Sodium tetrahydroborate (0.31 g, 8.1 mmol) was added to a mixture of1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone (from Example 11,Step 1) (1.5 g, 5.4 mmol) in methanol (25 mL) at 0° C. and the resultantreaction mixture was stirred at room temperature for 1 hour. The solventwas removed and the resulting residue was diluted with ethyl acetate,washed with sat. NaHCO₃, water, brine, then dried over Na₂SO₄, filteredand concentrated. The crude product was purified by silica gelchromatography, eluting with 0 to 40% EtOAc in hexanes (0.30 g, 90%).

Step 2.4-[3-Chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile

A mixture of 1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanol (0.30g, 1.1 mmol),4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carbonitrile(from Combi-Blocks, 0.27 g, 1.2 mmol), sodium carbonate (230 mg, 2.1mmol) and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II),complex with dichloromethane (1:1) (100 mg, 0.13 mmol) in acetonitrile(8 mL)/water (2 mL) was degassed and then refilled with N₂. The reactionwas stirred at 95° C. for 2 hours, then cooled and diluted with ethylacetate, washed with sat. NaHCO₃, water, brine, dried over Na₂SO₄,filtered and concentrated. The crude product was purified by silica gelchromatography, eluting with 0 to 40% EtOAc in hexanes (0.249 g, 75%).LCMS calculated for C₁₆H₁₆ClN₂O₂(M+H)⁺: m/z=303.1; Found: 303.0.

Step 3.4-[3-Chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile

A mixture of cyanuric chloride (170 mg, 0.94 mmol) andN,N-dimethylformamide (73 μL, 0.94 mmol) was stirred at room temperaturefor 10 minutes and then a solution of4-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile(190 mg, 0.628 mmol) in methylene chloride (4 mL) was added and thereaction was stirred at room temperature overnight. The mixture wasdiluted with methylene chloride, washed with sat. NaHCO₃, water, brine,dried over Na₂SO₄, filtered and concentrated. The crude product was useddirectly in the next step without purification (121 mg, 60%). LCMScalculated for C₁₆H₁₅Cl₂N₂O (M+H)⁺: m/z=321.0; Found: 321.0.

Step 4.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile

Sodium hydride (20 mg, 0.50 mmol) was added to a mixture of4-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile(90 mg, 0.28 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (63 mg,0.42 mmol) in N,N-dimethylformamide (4 mL) and the reaction was stirredet 30° C. overnight. The mixture was cooled, treated with water and thenfiltered to provide the desired product. LCMS calculated for C₂₂H₂₁ClN₇O(M+H)⁺: m/z=434.1; Found: 434.2.

Step 5.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carboxylicacid

Sodium hydroxide (1.0 M) in water (0.70 mL, 0.70 mmol) was added to amixture of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile(0.060 g, 0.14 mmol) in ethanol (1.0 mL) and the resultant mixture washeated at 95° C. for 6 hours. At this time, conc. HCl was added toadjust pH to ˜3. The solvent was removed and the residue was used in thenext step without further purification. LCMS calculated forC₂₂H₂₂ClN₆O₃(M+H)⁺: m/z=453.1; Found: 453.2.

Step 6.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide

2.0 M Dimethylamine in THF (0.14 mL, 0.28 mmol) was added to a solutionof4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carboxylicacid (9.6 mg, 0.021 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(10 mg, 0.03 mmol) in N,N-dimethylformamide (0.7 mL) at room temperaturefollowed by addition of triethylamine (8.8 μL, 0.064 mmol). The reactionwas stirred for 1 hour. The crude mixture was purified using RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 30 mL/min) to givethe desired product. The product was isolated as a racemic mixture. LCMScalculated for C₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2; Found: 480.2.

Example 17.4-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N-methylpicolinamide

This compound was prepared using procedures analogous to those forExample 16, Step 6, with 2.0 M solution of methylamine in THF replacing2.0 M dimethylamine in THF. The product was isolated as a racemicmixture. LCMS calculated for C₂₃H₂₅ClN₇O₂(M+H)⁺: m/z=466.2; Found:466.2.

Example 18.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)pyridine-2-carboxamide

This compound was prepared using procedures analogous to those forExample 16, Step 6, with ethanolamine replacing 2.0 M dimethylamine inTHF. The product was isolated as a racemic mixture. LCMS calculated forC₂₄H₂₇ClN₇O₃(M+H)⁺: m/z=496.2; Found: 496.2.

Example 19.4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide

This compound was prepared using procedures analogous to those forExample 16, Step 6, with 2-(methylamino)ethanol replacing 2.0 Mdimethylamine in THF. The product was isolated as a racemic mixture.LCMS calculated for C₂₅H₂₉ClN₇O₃(M+H)⁺: m/z=510.2; Found: 510.2.

Example 20.2-(4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-1H-pyrazol-1-yl)ethanol

Step 1. 3-Bromo-1-chloro-5-(1-chloroethyl)-4-methoxy-2-methylbenzene

A mixture of cyanuric chloride (1.7 g, 9.2 mmol) andN,N-dimethylformamide (710 μL, 9.2 mmol) was stirred at room temperaturefor 10 minutes and then a solution of1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanol (from Example 16,Step 1) (1.72 g, 6.15 mmol) in methylene chloride (34 mL) was added andthe reaction was stirred at room temperature overnight. The mixture wasdiluted with methylene chloride, washed with sat. NaHCO₃, water, brine,dried over Na₂SO₄, filtered and concentrated. The crude product waspurified by silica gel chromatography, eluting with 0 to 10% EtOAc inhexanes (1.01 g, 60%).

Step 2.1-[1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Sodium hydride (36 mg, 0.91 mmol) was added to a mixture of3-bromo-1-chloro-5-(1-chloroethyl)-4-methoxy-2-methylbenzene (150 mg,0.503 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (110 mg, 0.76mmol) in N,N-dimethylformamide (8 mL) and the reaction was stirred at30° C. overnight. The mixture was diluted with methylene chloride,washed with sat. NaHCO₃, water, brine, dried over Na₂SO₄, filtered andconcentrated. The crude product was purified by silica gelchromatography, eluting with 0 to 70% EtOAc in CH₂Cl₂ (103 mg, 50%).LCMS calculated for C₁₆H₁₈BrClN₅O (M+H)⁺: m/z=410.0; Found: 410. Theracemic products were applied on a Phenomenex Lux-Cellulose 1 column(21.1×250 mm, 5 micron particle size), eluting with 5% ethanol inhexanes at a flow rate of 18 mL/min, ˜13 mg/injection, to provide twoenantiomers.

Step 3.1-(2-{[tert-Butyl(dimethyl)silyl]oxy}ethyl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole

Potassium tert-butoxide (1.0 M) in THF (0.60 mL, 0.60 mmol) was added toa solution of4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (0.1 g, 0.5mmol) in N,N-dimethylformamide (1.5 mL) at 0° C. The reaction mixturewas stirred at room temperature for 5 minutes, then cooled to 0° C. andtreated with (2-bromoethoxy)(tert-butyl)dimethylsilane (0.2 mL, 0.8mmol). The reaction was stirred at room temperature overnight, thendiluted with ethyl acetate, washed with sat. NaHCO₃, water, brine, driedover Na₂SO₄, filtered and concentrated to provide the crude productwhich was purified by silica gel chromatography eluting with 0 to 30%EtOAc in hexanes. Calculated for C₁₇H₃₄BN₂O₃Si (M+H)⁺: m/z=353.2; Found:353.1.

Step 4.2-(4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-1H-pyrazol-1-yl)ethanol

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(0.026 g, 0.062 mmol) (chiral pure, first peak from Step 2),1-(2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole(0.024 g, 0.069 mmol), sodium carbonate (13 mg, 0.12 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (6.1 mg, 0.0075 mmol) in acetonitrile (0.5mL)/water (0.1 mL) was degassed and then refilled with N₂. The reactionmixture was stirred at 95° C. for 2 hours, then treated with conc. HCl(0.1 mL) and then stirred at room temperature for 1 hour. The crudemixture was purified using RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 30 mL/min) to give the desired product. The product wasisolated as a single enantiomer. LCMS calculated for C₂₁H₂₅ClN₇O₂(M+H)⁺:m/z=442.2; Found: 442.2.

Example 21.3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-N,N,6′-trimethylbiphenyl-4-carboxamidetrifluoroacetate

Step 1. Methyl3′-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5′-chloro-3-fluoro-2′-methoxy-6′-methylbiphenyl-4-carboxylate

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(60 mg, 0.15 mmol, chiral pure, first peak from Example 20, Step 2),[3-fluoro-4-(methoxycarbonyl)phenyl]boronic acid (from Combi-Blocks,0.041 g, 0.20 mmol), sodium carbonate (36 mg, 0.34 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (6 mg, 0.007 mmol) in acetonitrile (1.2mL)/water (0.3 mL) was vacuumed and then refilled with N₂. The reactionwas stirred at 95° C. for 2 hours. Then solvent was removed and thecrude mixture was purified by silica gel chromatography, eluting with 0to 70% EtOAc in CH₂Cl₂, to give the desired product (54 mg, 75%). LCMScalculated for C₂₄H₂₄ClFN₅O₃ (M+H)⁺: m/z=484.2; Found: 484.1.

Step 2.3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-6′-methylbiphenyl-4-carboxylicacid

Lithium hydroxide, monohydrate (13 mg, 0.31 mmol) was added to asolution of methyl3′-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-6′-methylbiphenyl-4-carboxylatemade above (0.030 g, 0.062 mmol) in methanol (0.2 mL)/tetrahydrofuran(0.2 mL)/water (0.09 mL). The reaction was stirred at room temperaturefor 1.5 h, then treated with conc. HCl (60 uL) to adjust pH to 2. Thesolvent was removed to provide the crude product which was used in nextstep without further purification. LCMS calculated for C₂₃H₂₂ClFN₅O₃(M+H)⁺: m/z=470.1; Found: 470.2.

Step 3.3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-N,N,6′-trimethylbiphenyl-4-carboxamide trifluoroacetate

2.0 M Dimethylamine in THF (0.1 mL, 0.2 mmol) was added to a solution of3′-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-6′-methylbiphenyl-4-carboxylicacid (12 mg, 0.026 mmol) made above andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(20 mg, 0.04 mmol) in N,N-dimethylformamide (0.7 mL) at room temperaturefollowed by addition of triethylamine (11 μL, 0.077 mmol). The reactionwas stirred for 1 hour, quenched with water. The crude mixture wasapplied on RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as TFA salt. The product was isolated as asingle enantiomer. LCMS calculated for C₂₅H₂₇ClFN₆O₂(M+H)⁺: m/z=497.2;Found: 497.2.

Example 22.3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-N,6′-dimethylbiphenyl-4-carboxamidetrifluoroacetate

This compound was prepared using procedures analogous to those forExample 21, Step 3, with 2.0 M methylamine in THF replacing 2.0 Mdimethylamine in THF. The product was isolated as a single enantiomer.LCMS calculated for C₂₄H₂₅ClFN₆O₂(M+H)⁺: m/z=483.2; Found: 483.2.

Example 23.5-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N-(2-hydroxyethyl)picolinamidetrifluoroacetate

Step 1.5-[3-Chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile

A mixture of 1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanol (0.15g, 0.54 mmol),5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carbonitrile(from Frontier, 0.14 g, 0.59 mmol), sodium carbonate (110 mg, 1.1 mmol)and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (52 mg, 0.064 mmol) in acetonitrile (4mL)/water (1 mL) was degassed and then refilled with N₂. The reactionwas stirred at 95° C. for 2 h, cooled, diluted with ethyl acetate,washed with sat. NaHCO₃, water, brine, and then dried over Na₂SO₄,filtered and concentrated. The crude product was purified by silica gelchromatography, eluting with 0 to 40% EtOAc in hexanes, to give thedesired product (114 mg, 70%). LCMS calculated for C₁₆H₁₆ClN₂O₂(M+H)⁺:m/z=303.1; Found: 303.0.

Step 2.5-[3-Chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile

A mixture of cyanuric chloride (170 mg, 0.94 mmol) andN,N-dimethylformamide (73 μL, 0.94 mmol) was stirred at room temperaturefor 10 minutes and then a solution of5-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile(190 mg, 0.628 mmol) in methylene chloride (4 mL) was added and thereaction was stirred at room temperature overnight. The mixture wasdiluted with methylene chloride, washed with sat. NaHCO₃, water, brine,dried over Na₂SO₄, then filtered and concentrated. The resultant crudeproduct was used directly in the next step without further purification(110 mg, 55%). LCMS calculated for C₁₆H₁₅Cl₂N₂O (M+H)⁺: m/z=321.0;Found: 321.0.

Step 3.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile

Sodium hydride (20 mg, 0.50 mmol) was added to a mixture of5-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]pyridine-2-carbonitrile(90 mg, 0.28 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (63 mg,0.42 mmol) in N,N-dimethylformamide (4 mL) and the reaction was stirredat 30° C. overnight. The mixture was treated with water and thenfiltered to provide the desired product. LCMS calculated for C₂₂H₂₁ClN₇O(M+H)⁺: m/z=434.1; Found: 434.2.

Step 4.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carboxylicacid

Sodium hydroxide (1.0 M) in water (0.70 mL, 0.70 mmol) was added to amixture of5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile(0.060 g, 0.14 mmol) in ethanol (1.0 mL). The reaction was heated at 95°C. for 6 hours, followed by the addition of conc. HCl to adjust pH to˜3. The solvent was removed and the resultant residue was used in thenext step without further purification. LCMS calculated forC₂₂H₂₂ClN₆O₃(M+H)⁺: m/z=453.1; Found: 453.2.

Step 5.5-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N-(2-hydroxyethyl)picolinamidetrifluoroacetate

Ethanolamine (15 μL, 0.25 mmol) was added to a solution of5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carboxylicacid (9.6 mg, 0.021 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(10 mg, 0.03 mmol) in N,N-dimethylformamide (0.7 mL) at room temperaturefollowed by addition of triethylamine (8.8 μL, 0.064 mmol). The reactionwas stirred for 1 hour, and then quenched with water. The crude mixturewas applied on RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as TFA salt. The product was isolated as aracemic mixture. LCMS calculated for C₂₄H₂₇ClN₇O₃ (M+H)⁺: m/z=496.2;Found: 496.2.

Example 24.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamidetrifluoroacetate

This compound was prepared using procedures analogous to those forExample 23, with 2-(methylamino)ethanol replacing ethanolamine. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₅H₂₉ClN₇O₃ (M+H)⁺: m/z=510.2; Found: 510.2.

Example 40.4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-cyano-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide

Catalyst preformation: Anhydrous dimethylacetamide (DMA) was purged witha gentle stream of N₂ for 30 minutes prior to use. A 50 mM solution ofH₂SO₄ was prepared with 10 mL dimethylacetamide and 26.8 μL of conc.H₂SO₄ and then purged with N₂ for 10 minutes. To an 8 mL vial equippedwith a magnetic stir bar and septum cap were added Pd(OAc)₂ (22.5 mg,100 μmol) and 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl(95.3 mg, 200 μmol). The vial was evacuated and filled with N₂ threetimes, purged with a gentle stream of N₂ for 10 minutes. H₂SO₄ (2.0 mL,50 mM in DMA) was added, and the catalyst mixture was stirred in an oilbath at 80° C. for 30 minutes to give a homogeneous coffee-brownsolution.

The above catalyst (0.05 mL) was added to a mixture of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide(from Example 19) (4.0 mg, 0.0078 mmol), zinc (0.22 mg, 0.0034 mmol) andzinc cyanide (0.92 mg, 0.0078 mmol) in N,N-dimethylacetamide (0.1 mL).The mixture was degassed and then the reaction was heated at 120° C. for1.5 hours. The crude mixture was applied on RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₆H₂₉N₈O₃ (M+H)⁺: m/z=501.2; Found: 501.2.

Example 41.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

Step 1:N-(2,4-Dimethoxybenzyl)-3-methyl-1H-pyrazolo[4,3-c]pyridin-4-amine

A solution of 4-chloro-3-methyl-1H-pyrazolo[4,3-c]pyridine (330 mg, 1.9mmol) and 1-(2,4-dimethoxyphenyl)methanamine (0.58 mL, 3.9 mmol) in1-butanol was heated in the microwave at 150° C. for 40 minutes.Purification via preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) gave the desired product (240 mg, 42%). LCMS forC₁₆H₁₉N₄O₂ (M+H)⁺: m/z=299.1; Found: 299.2.

Step 2:5-[3-Chloro-5-(1-{4-[(2,4-dimethoxybenzyl)amino]-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl}ethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide

A solution ofN-(2,4-dimethoxybenzyl)-3-methyl-1H-pyrazolo[4,3-c]pyridin-4-amine (110mg, 0.37 mmol) in N,N-dimethylformamide (2 mL) was treated with sodiumhydride (30 mg, 0.75 mmol) and stirred at 20° C. for 30 minutes. Thereaction mixture was treated with a solution of5-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(130 mg, 0.34 mmol) in N,N-dimethylformamide (1 mL) and heated at 50° C.overnight. The reaction mixture was diluted with water and extractedwith ethyl acetate (2×). The combined organic extracts were washed withwater and brine, dried with magnesium sulfate, filtered, andconcentrated to a crude residue. Purification via preparative LCMS(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 60 mL/min) gave thedesired product (110 mg, 49%). LCMS for C₃₄H₃₈ClN₆O₄ (M+H)⁺: m/z=629.3;Found: 629.1.

Step 3:5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

A solution of5-[3-chloro-5-(1-{4-[(2,4-dimethoxybenzyl)amino]-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl}ethyl)-6-methoxy-2-methylphenyl]-N,N-dimethylpyridine-2-carboxamide(85 mg, 0.14 mmol) in methylene chloride (2 mL) was treated withtrifluoroacetic acid (2 mL) and stirred at 20° C. for 3 hours and at 40°C. for 20 minutes. Purification via preparative LCMS (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%trifluoroacetic acid, at flow rate of 60 mL/min) gave the desiredproduct (44 mg, 46%). The product was isolated as a racemic mixture.LCMS for C₂₅H₂₈ClN₆O₂(M+H)⁺: m/z=479.2; Found: 479.0. ¹H NMR (300 MHz,DMSO-d₆): δ 12.8 (br s, 0.5H), 8.50 (br s, 0.5H), 8.37 (br s, 2H),7.91-7.86 (m, 0.5H), 7.80-7.75 (m, 0.5H), 7.68-7.58 (m, 3H), 7.17 (d,J=7.3 Hz, 1H), 6.19 (q, J=6.9 Hz, 1H), 3.04 (s, 3H), 3.01 (s, 3H), 2.94(s, 3H), 2.61 (s, 3H), 2.05 (s, 3H), 1.83 (d, J=6.9 Hz, 3H).

Example 43.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[5-(methylsulfonyl)pyridin-3-yl]benzonitrile

Step 1. 1-(3-bromo-5-chloro-4-fluoro-2-hydroxyphenyl)ethanone

1-(5-Chloro-4-fluoro-2-hydroxyphenyl)ethanone (e.g., from Example 13,step 1) (20.0 g, 101 mmol, 1.00 eq) and a 50% aqueous sulfuric acid (120mL) were added to the flask. The resulting mixture was heated to 60° C.in a water bath with stirring. N-Bromosuccinimide (21.52 g, 120.9 mmol,1.20 eq) was added in three portions [7.0 g+7.0 g+7.52 g] in 8 minuteintervals. After the reaction mixture was heated at 60° C. for 3 hours,the reaction was complete. The reaction mixture was diluted with water(160 ml) and dichloromethane (DCM) (300 ml), and the mixture was stirredfor 0.5 hour. The organic layer was separated and the aqueous layer wasextracted with dichloromethane (100 ml). The combined organic layerswere washed with 1 N HCl (100 ml×2), water (100 ml), brine (60 ml), andconcentrated under reduced pressure to afford the crude product (29.1 g)as a yellowish solid. The crude product was dissolved in HOAc (100 ml)and then diluted with water (200 ml) under stirring. The resultingmixture was stirred for 20 min at room temperature and the product wascollected by filtration and dried to give1-(3-bromo-5-chloro-4-fluoro-2-hydroxyphenyl)ethanone (21.8 g, 80.9%) asa yellowish solid. ¹H-NMR (300 MHz, CDCl₃) δ 13.18 (s, 1H, —OH), 7.78(d, J=7.78 Hz, 1H), 2.63 (s, 3H).

Step 2. 4-Acetyl-2-bromo-6-chloro-3-ethoxybenzonitrile

1-(3-Bromo-5-chloro-4-fluoro-2-hydroxyphenyl)ethanone (2.0 g, 7.5 mmol)was combined with potassium cyanide (0.58 g, 9.0 mmol) inN,N-dimethylformamide (16 mL, 210 mmol) and heated to 85° C. in an oilbath. After heating for 18 hours, the reaction was allowed to cool toroom temperature and iodoethane (0.90 mL, 11 mmol) and potassiumcarbonate (2.1 g, 15 mmol) were added. The reaction was heated to 65° C.and monitored by LC/MS. After heating for 3 hours the reaction wascomplete and allowed to cool to room temperature, then taken up in ethylacetate and washed with water, brine, and dried over magnesium sulfate.The resultant solution was concentrated to give the crude product as adark oil. The product was purified by flash column chromatography onsilica gel eluting hexane: ethyl acetate gradient to give4-acetyl-2-bromo-6-chloro-3-ethoxybenzonitrile (1.15 gm, 50%) as a solidresidue, LCMS calculated for C₁₁H₉BrClNO₂(M+H)⁺: m/z=301.9, 303.9;found: (no ionization)

Step 3. 2-Bromo-6-chloro-3-ethoxy-4-(1-hydroxyethyl)benzonitrile

Sodium tetrahydroborate (38 mg, 0.99 mmol) was added to a mixture of4-acetyl-2-bromo-6-chloro-3-ethoxybenzonitrile (200 mg, 0.7 mmol) inmethanol (5 mL, 100 mmol) at 0° C. The reaction was stirred at roomtemperature for 1 hour, concentrated and partitioned between water andEtOAc. The combined organic layers were washed with brine, dried overMgSO₄, filtered and concentrated to give crude2-bromo-6-chloro-3-ethoxy-4-(1-hydroxyethyl)benzonitrile as a clear oil(0.15 gm, 100%), LCMS calculated for CuIHiiBrClNO₂(M+H)⁺: m/z=303.9,305.9; found: 304.0, 305.9.

Step 4. 2-Bromo-6-chloro-4-(1-chloroethyl)-3-ethoxybenzonitrile

Cyanuric chloride (0.11 g, 0.59 mmol) was dissolved inN,N-dimethylformamide (3 mL, 40 mmol). After stirring for a few minutes,a solution of 2-bromo-6-chloro-3-ethoxy-4-(1-hydroxyethyl)benzonitrile(150 mg, 0.49 mmol) in methylene chloride (3 mL, 50 mmol) was added. Theresulting mixture was stirred at room temperature overnight. Thereaction was partitioned between water and dichloromethane. The organiclayer was washed with sat. NaHCO₃ solution, water, brine, dried overMgSO₄, and concentrated. The crude product was purified by flash columnchromatography, eluting a gradient of 0-30% EtOAc/Hexane to give2-bromo-6-chloro-4-(1-chloroethyl)-3-ethoxybenzonitrile (0.12 gm, 75%)as a semisolid, LCMS calculated for C₁₁H₁₀BrCl₂NO (M+H)⁺: m/z=323.9,320.9; found: (poor ionization).

Step 5.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-bromo-6-chloro-3-ethoxybenzonitrile

Sodium hydride (16 mg, 0.41 mmol) was added to a mixture of3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (33 mg, 0.22 mmol) inN,N-dimethylformamide (3 mL, 40 mmol) and was stirred for 10 minutes.2-bromo-6-chloro-4-(1-chloroethyl)-3-ethoxybenzonitrile (60 mg, 0.2mmol) in N,N-dimethylformamide (2 mL) was added and the reaction wasstirred at 50° C. overnight. The mixture was diluted with methylenechloride, washed with sat'd NaHCO₃, water, brine, dried over Na₂SO₄,filtered and concentrated. The product was purified by flash columnchromatography eluting with CH₂Cl₂/MeOH 0-10%, to give4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-bromo-6-chloro-3-ethoxybenzonitrile(0.05 gm, 60%) as a solid, LCMS calculated for C₇H₆BrClN₆O (M+H)⁺:m/z=437.0, 435.0; found: 436.9, 434.7.

Step 6.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[5-(methylsulfonyl)pyridin-3-yl]benzonitrile

To a mixture of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-bromo-6-chloro-3-ethoxybenzonitrile(20 mg, 0.04 mmol) and3-(methylsulfonyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine(19 mg, 0.069 mmol) in acetonitrile (2 mL, 40 mmol) was added sodiumcarbonate (10 mg, 0.09 mmol) in water (0.5 mL, 30 mmol). The reactionwas degassed with bubbling nitrogen.[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II) complex withdichloromethane (1:1) (2 mg, 0.002 mmol) was added and degassed morewith N₂. Reaction was heated at 100° C. for 2 hours. The crude productwas purified on preparative LC-MS (acetonitrile, water, TFA) to give thedesired product (0.004 g, 20%) as white amorphous solid. The product wasisolated as a racemic mixture. LCMS calculated for C₂₃H₂₂ClN₇O₃S (M+H)⁺:m/z=512.1; found: 512.2. ¹H NMR (500 MHz, DMSO) δ 9.20 (d, J=2.1 Hz,1H), 9.12 (d, J=1.9 Hz, 1H), 8.61 (t, J=2.0 Hz, 1H), 8.12 (s, 1H), 7.80(s, 1H), 6.36 (q, J=7.0 Hz, 1H), 3.54 (dt, J=14.0, 7.0 Hz, 1H), 3.37 (s,3H), 3.36-3.30 (m, 1H), 2.58 (s, 3H), 1.81 (d, J=7.0 Hz, 3H), 0.92 (t,J=6.9 Hz, 3H).

Example 44.5-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-6-cyano-2-ethoxyphenyl)-N,N-dimethylpicolinamide

The title compound was prepared in analogous manor as Example 43, step 6but usingN,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)picolinamide(Peptech, Cat#BE1622) to give the crude product which was purified onpreparative LC-MS (acetonitrile, water, TFA) to give the desired product(0.005 g, 22%) as white amorphous solid. The product was isolated as aracemic mixture. LCMS calculated for C₂₅H₂₅ClN₈O₂(M+H)⁺: m/z=505.1;found: 505.1. ¹H NMR (500 MHz, DMSO) δ 8.72 (dd, J=2.1, 0.7 Hz, 1H),8.14-8.12 (m, 1H), 8.11 (s, 1H), 7.75 (s, 1H), 7.71 (dd, J=8.0, 0.7 Hz,1H), 6.35 (q, J=7.0 Hz, 1H), 3.61-3.48 (m, 1H), 3.42-3.31 (m, 1H), 3.03(s, 3H), 2.95 (s, 3H), 2.57 (s, 3H), 1.80 (d, J=7.1 Hz, 3H), 0.92 (t,J=7.0 Hz, 3H).

Example 65.5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylnicotinamide

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(25 mg, 0.061 mmol) (chiral pure, first peak from Example 20, Step 2),N,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)nicotinamide(from PepTech) (25 mg, 0.091 mmol), sodium carbonate (13 mg, 0.12 mmol)and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium (II),complex with dichloromethane (1:1) (9.9 mg, 0.012 mmol) in acetonitrile(0.8 mL)/water (0.3 mL) was degassed with N₂ and then stirred at 95° C.for 2 h. The mixture was filtered and the filtrate purified by RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 30 mL/min) to givethe desired product. The product was isolated as a single enantiomer.LCMS calculated for C₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2; Found: 480.2. ¹H NMR(500 MHz, DMSO-d₆) δ 8.64 (1H, s), 8.54 (1H, br s), 8.13 (1H, s), 7.82(1H, m), 7.53 (1H, s), 7.42 (2H, br s), 6.28 (1H, q, J=6.5 Hz), 3.22(3H, s), 2.95 (6H, m), 2.58 (3H, s), 2.04 (3H, s), 1.77 (3H, d, J=6.5Hz) ppm.

Example 66.5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamidebis(trifluoroacetate)

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(25 mg, 0.061 mmol) (chiral pure, first peak from Example 20, Step 2),N,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carboxamide(25 mg, 0.091 mmol), sodium carbonate (13 mg, 0.12 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]-dichloropalladium (II), complexwith dichloromethane (1:1) (9.9 mg, 0.012 mmol) in acetonitrile (0.8mL)/water (0.3 mL) was degassed with N₂ and then stirred at 95° C. for 2hours. After cooling to room temperature, the mixture was filtered andthe filtrate purified on RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.05% trifluoroacetic acid, atflow rate of 30 mL/min) to give the desired product as bis-TFA salt. Theproduct was isolated as a single enantiomer. LCMS calculated forC₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2; Found: 480.2. ¹H NMR (500 MHz, DMSO-d₆)δ: 8.78 (2H, br s), 8.48 (1H, m), 8.36 (1H, s), 7.86 (1H, br s), 7.65(1H, br s), 7.58 (1H, s), 6.33 (1H, q, J=7.0 Hz), 3.19 (3H, s), 3.03(3H, s), 2.97 (3H, s), 2.62 (3H, s), 2.06 (3H, s), 1.81 (3H, d, J=7.0Hz) ppm.

Example 67.1-{1-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Step 1.1-[1-(3-Azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidine-1-carboxylate(1.6 g, 3.2 mmol, from Example 13, Step 7) was treated with 4.0 Mhydrogen chloride in dioxane (8.15 mL, 32.6 mmol) in methylene chloride(17 mL) at room temperature for 2 h. The mixture was concentrated todryness to give the desired product. LCMS calculated for C₁₈H₂₁ClFN₆O(M+H)⁺: m/z=391.1; Found: 391.1.

Step 2.1-{-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (0.90 g, 1.9 mmol, Example 67, step 1), acetone (1.0 mL,14 mmol) and triethylamine (2.5 mL, 18 mmol) in methylene chloride (20mL) was added sodium triacetoxyborohydride resin (2.5 g, 5.8 mmol). Themixture was stirred at room temperature for 2 h, then filtered, washedwith water, dried over MgSO₄, filtered and concentrated to give crudeproduct (870 mg, 100%). LCMS calculated for C₂₁H₂₇ClFN₆O (M+H)⁺:m/z=433.2; Found: 433.1.

Step 3. Single Enantiomer of1-{1-[5-chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Enantiomers of1-{1-[5-chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(870 mg, 2.0 mmol) were separated on a Phenomenex Lux Cellulose-2column, eluting with 10% ethanol in hexanes, at flow rate of 18 mL/min,and column loading of ˜8 mg/injection to separate two enantiomers. Firstpeak retention time 10.9 min; second peak retention time 13.6 min. Thefractions of the 1st peak (110 mg, 13%) were concentrated and purifiedusing RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product. The product was isolated as asingle enantiomer. LCMS calculated for C₂₁H₂₇ClFN₆O (M+H)⁺: m/z=433.2;Found: 433.1.

Example 68.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidin-1-yl)propan-2-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (15 mg, 0.032 mmol, from Example 67, Step 1) andtriethylamine (18 μL, 0.13 mmol) in ethanol (0.53 mL) was added(S)-(−)-methyloxirane (6.8 μL, 0.097 mmol). The resulting mixture washeated at 90° C. for 3 h, then purified on RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product. Theenantiomers were separated on a Phenomenex Lux Cellulose C-4 column (5μM, 21.2×250 mm), eluting with 20% ethanol in hexanes, at flow rate of18 mL/min, to give two enantiomers. First peak (2.7 mg, 18%) retentiontime 8.9 min; LCMS calculated for C₂₁H₂₇ClFN₆O₂(M+H)⁺: m/z=449.2; Found:449.1. ¹H NMR (DMSO-d₆, 500 MHz) δ 8.11 (1H, s), 7.42 (1H, d, J=8.5 Hz),7.25 (2H, br s), 6.21 (1H, q, J=7.5 Hz), 4.28 (1H, d, J=4.0 Hz), 3.82(3H, m), 3.62 (3H, s), 3.55 (1H, m), 3.05 (1H, m), 2.97 (1H, m), 2.55(3H, s), 2.28 (2H, m), 1.70 (2H, d, J=7.5 Hz), 1.00 (3H, d, J=6.0 Hz)ppm. Second peak retention time 10.0 min.

Example 71.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidin-1-yl)ethanol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (19 mg, 0.041 mmol, racemic intermediate from Example67, Step 1) and triethylamine (28 μL, 0.20 mmol) in methanol (0.1mL)/acetonitrile (0.1 mL)/tetrahydrofuran (0.1 mL) was added{[tert-butyl(dimethyl)silyl]oxy}acetaldehyde (39 μL, 0.20 mmol),followed by sodium triacetoxyborohydride (22 mg, 0.10 mmol). Theresulting mixture was stirred overnight at room temperature. The mixturewas treated with 6.0 M hydrogen chloride in water (0.07 mL, 0.4 mmol) atroom temperature for 10 min and then purified on RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 30 mL/min) to give the desiredproduct (2.5 mg, 13%). The product was isolated as a racemic mixture.LCMS calculated for C₂₀H₂₅ClFN₆O₂(M+H)⁺: m/z=435.2; Found: 435.1.

Example 72.1-{1-[5-Chloro-4-fluoro-2-methoxy-3-(1-oxetan-3-ylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-4-fluoro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (19 mg, 0.041 mmol racemic intermediate from Example 67,Step 1) and triethylamine (28 μL, 0.20 mmol) in methanol (0.1mL)/acetonitrile (0.1 mL)/tetrahydrofuran (0.1 mL) was added 37%formaldehyde (15 μL, 0.20 mmol), followed by sodiumtriacetoxyborohydride (22 mg, 0.10 mmol). The resulting mixture wasstirred overnight at room temperature. The mixture was purified onRP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product (1.2 mg, 6.3%). The product wasisolated as a racemic mixture. LCMS calculated for C₁₉H₂₃ClFN₆O (M+H)⁺:m/z=405.2; Found: 405.1.

Example 94.1-{1-[5-Chloro-2-ethoxy-3-(1-isopropylazetidin-3-yl)-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

Step 1. Benzyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidine-1-carboxylate

Cyanuric chloride (200 mg, 1.1 mmol) was added to N,N-dimethylformamide(0.083 mL, 1.1 mmol) at room temperature. After the formation of a whitesolid (ca. 10 minutes), methylene chloride (5 mL) was added, followed bybenzyl3-[3-chloro-6-ethoxy-5-(1-hydroxyethyl)-2-methylphenyl]azetidine-1-carboxylate(310 mg, 0.77 mmol). After addition, the resultant mixture was stirredat room temperature overnight. Water was added, and then diluted withdichloromethane. The organic phases were washed with sat. NaHCO₃solution, water and brine, dried over MgSO₄, concentrated and purifiedon silica gel (eluting with 0 to 40% EtOAc/hexanes) to give the desiredproduct (140 mg, 43%). LCMS calculated for C₂₂H₂₆Cl₂NO₃ (M+H)⁺:m/z=422.1; Found: 422.0.

A mixture of benzyl3-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-methylphenyl]azetidine-1-carboxylate(0.375 g, 0.888 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(0.16 g, 1.1 mmol), cesium carbonate (0.43 g, 1.3 mmol) and potassiumiodide (15 mg, 0.089 mmol) in N,N-dimethylformamide (2.8 mL) was heatedat 140° C. for 1 h. The mixture was diluted with ether, and washed withwater. The organic layers were concentrated and purified on silica gel(eluting with 0 to 100% EtOAc in hexanes) to give the desired product(0.24 g, 50%). LCMS calculated for C₂₈H₃₂ClN₆O₃(M+H)⁺: m/z=535.2; Found:535.0. The enantionmers were separated on a Phenomenex Lux Cellulose C-2column (5 μM, 21.2×250 mm), eluting with 20% ethanol in hexanes, at flowrate of 18 mL/min, and column loading of ˜4.5 mg/injection to separatetwo enantiomers. First peak retention time: 21.2 min; second peakretention time: 24.6 min.

Step 2.1-[1-(3-Azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Benzyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidine-1-carboxylate(170 mg, 0.32 mmol, racemic intermediate) and 5% palladium (80 mg) werecombined in methanol (12 mL), to which was added 0.25 M hydrogenchloride in water (3.2 mL, 0.79 mmol). The suspension was hydrogenatedunder balloon pressure of H₂ at room temperature for 2 h. The suspensionwas filtered. The filtrate was neutralized with sat. NaHCO₃ solution,and extracted with dichloromethane. The combined organic layers weredried over MgSO₄ and filtered, concentrated to give the desired product(117 mg, 92%). LCMS calculated for C₂₀H₂₆ClN₆O (M+H)⁺: m/z=401.2; Found:401.1.

Step 3.1-{1-[5-Chloro-2-ethoxy-3-(1-isopropylazetidin-3-yl)-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminebis(trifluoroacetate)

Acetone (9.3 μL, 0.13 mmol) was added to1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(10.2 mg, 0.0254 mmol) in methanol (0.1 mL)/tetrahydrofuran (0.1mL)/acetonitrile (0.1 mL) and the mixture was stirred at roomtemperature for 10 min, before the addition of sodiumtriacetoxyborohydride (16 mg, 0.076 mmol). The reaction mixture wasstirred at room temperature for 4 h and then purified on RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.05% TFA, at flow rate of 30 mL/min) to give the desiredproduct as TFA salt (2.3 mg, 22%). The product was isolated as a singleenantiomer. LCMS calculated for C₂₃H₃₂ClN₆O (M+H)⁺: m/z=443.2; Found:443.1.

Example 95.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)ethanolbis(trifluoroacetate)

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(7.9 mg, 0.020 mmol, racemic intermediate from Example 94, Step 2) intetrahydrofuran (0.09 mL)/acetonitrile (0.09 mL)/methanol (0.09 mL) wasadded {[tert-butyl(dimethyl)silyl]oxy}acetaldehyde (19 μL, 0.098 mmol)and the mixture was stirred for 10 min before the addition of sodiumtriacetoxyborohydride (12 mg, 0.059 mmol). The resulting mixture wasstirred at room temperature for 4 h, then treated with 6.0 M hydrogenchloride in water (30 μL, 0.2 mmol) for 10 min. The mixture was purifiedon RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as TFA salt (3.2 mg, 40%). The product wasisolated as a racemic mixture. LCMS calculated for C₂₂H₃₀ClN₆O₂(M+H)⁺:m/z=445.2; Found: 445.1.

Example 96.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)propan-2-olbis(trifluoroacetate)

Step 1. Benzyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidine-1-carboxylate

The enantionmers from Example 94, Step 1 were separated on a PhenomenexLux Cellulose C-2 column (5 μM, 21.2×250 mm), eluting with 20% ethanolin hexanes, at flow rate of 18 mL/min, and column loading of ˜4.5mg/injection to separate two enantiomers. First peak retention time:21.2 min; second peak retention time: 24.6 min.

Step 2.1-[1-(3-Azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Benzyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidine-1-carboxylate(chiral intermediate from first peak of previous step) was hydrogenatedin the presence of 5% palladium as described in Example 94, Step 2 togive the desired chiral product. LCMS calculated for C₂₀H₂₆ClN₆O (M+H)⁺:m/z=401.2; Found: 401.1.

Step 3.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)propan-2-olbis(trifluoroacetate)

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(10 mg, 0.02 mmol, chiral intermediate from step 2) and triethylamine (9μL, 0.07 mmol) in isopropyl alcohol (0.05 mL) was added(S)-(−)-methyloxirane (4.5 μL, 0.064 mmol). The resulting mixture wasstirred at 90° C. overnight, cooled and purified on RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.05%TFA, at flow rate of 30 mL/min) to give the desired product as TFA salt(3.4 mg, 34%). The product was isolated as a single diastereomer. LCMScalculated for C₂₃H₃₂ClN₆O₂(M+H)⁺: m/z=459.2; Found: 459.1.

Example 99.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-oltrifluoroacetate

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(9.8 mg, 0.024 mmol, racemic intermediate from Example 94, Step 2),N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uroniumhexafluorophosphate (14 mg, 0.037 mmol) and triethylamine (10 μL, 0.073mmol) in N,N-dimethylformamide (0.15 mL) was added 85%(2S)-2-hydroxypropanoic acid in water (3.2 μL, 0.037 mmol). Theresulting mixture was stirred for 2 h at room temperature. The mixturewas purified on RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.05% TFA, at flow rate of 30 mL/min) togive the desired product as trifluoroacetic acid (TFA) salt (2.9 mg,29%). The product was isolated as a racemic mixture. LCMS calculated forC₂₃H₃₀ClN₆O₃(M+H)⁺: m/z=473.2; Found: 473.1.

Example 102.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propan-2-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (21 mg, 0.046 mmol) (Example 1, step 7, chiralintermediate from peak 1) and triethylamine (20 μL, 0.1 mmol) inisopropyl alcohol (0.10 mL) was added (S)-(−)-methyloxirane (3.2 μL,0.046 mmol). The resulting mixture was stirred at 90° C. After 90 min,additional (S)-(−)-methyloxirane (6.4 uL) was added and stirred at 90°C. overnight. After cooling, the mixture was diluted with methanol andpurified using RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give 6 mg (30%) of the product. The product was isolatedas a single diastereomer. LCMS calculated for C₂₂H₃₀ClN₆O₂(M+H)⁺:m/z=445.2; Found: 445.2.

Example 104.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)ethanol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (20 mg, 0.04 mmol) (Example 1, step 7, chiralintermediate from peak 1), {[tert-butyl(dimethyl)silyl]oxy}acetaldehyde(8.3 mg, 0.048 mmol), and triethylamine (19 μL, 0.14 mmol) in methylenechloride (0.3 mL) was added sodium triacetoxyborohydride resin (38 mg,0.087 mmol). The resulting mixture was stirred overnight at roomtemperature. The mixture was filtered and concentrated. The crudeproduct was dissolved in tetrahydrofuran (1 mL) and cooled to 0° C. 1.0M Tetra-n-butylammonium fluoride in THF (0.44 mL, 0.44 mmol) was addedand warmed to room temperature. After 3 h, the solvents were evaporated.The crude was purified using RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 30 mL/min) to give 8.1 mg (40%) of the desired product. Theproduct was isolated as a single enantiomer. LCMS calculated forC₂₁H₂₈ClN₆O₂(M+H)⁺: m/z=431.2; Found: 431.3.

Example 105.(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)acetonitrile

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (16 mg, 0.035 mmol, chiral intermediate from peak 1 ofExample 1, Step 7) and triethylamine (14 μL, 0.10 mmol) in acetonitrile(0.7 mL) was added bromoacetonitrile (2.7 μL, 0.038 mmol). The resultingmixture was stirred at room temperature for 2.5 h. The mixture wasdiluted with acetonitrile and purified by using RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.05%TFA, at flow rate of 30 mL/min) to give the desired product as the TFAsalt. The pure fractions were partially evaporated and then made basicby the addition of 1 N NaOH. The aqueous mixture was extracted withdichloromethane (2×). The extracts were dried (MgSO4), filtered, andconcentrated. The solid was dried in vacuo to give 6.9 mg (46%) of thedesired product. The product was isolated as a single enantiomer. LCMScalculated for C₂₁H₂₅ClN₇O (M+H)⁺: m/z=426.2; Found: 426.0.

Example 108.1-(1-{5-Chloro-2-methoxy-4-methyl-3-[1-(2,2,2-trifluoroethyl)azetidin-3-yl]phenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (15 mg, 0.024 mmol, chrial intermediate from first peakof Example 1, step 7), 2,2,2-trifluoroethyl trifluoromethanesulfonate(6.8 mg, 0.029 mmol) and triethylamine (12 μL, 0.085 mmol) in methylenechloride (0.3 mL) was stirred over a weekend at room temperature. Thesolvents were evaporated and the crude purified using RP-HPLC (XBridgeC18 column, eluting with a gradient of acetonitrile/water containing0.1% ammonium hydroxide, at flow rate of 30 mL/min) to give 4.5 mg (39%)of the desired product. The product was isolated as a single enantiomer.LCMS calculated for C₂₁H₂₅ClF₃N₆ (M+H)⁺: m/z=469.2; Found: 469.1.

Example 110.(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-N-methylpropanamidetrifluoro acetate

A mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (26 mg, 0.067 mmol, chrial intermediate from peak 1 ofExample 1, Step 7), (2R)-2-bromopropanoic acid (7.3 μL, 0.081 mmol) andtriethylamine (19 μL, 0.13 mmol) in acetonitrile (0.8 mL) was stirredovernight at room temperature. The reaction was not complete so it washeated to 50° C. After 4 h, the solvents were evaporated. To the cruderesidue was added methylammonium chloride (4.5 mg, 0.067 mmol),N,N-dimethylformamide (0.2 mL), triethylamine (19 μL, 0.13 mmol), andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(45 mg, 0.10 mmol). The resulting mixture was stirred overnight at roomtemperature. The reaction mixture was added to a vial containing sat.NaHCO₃ and extracted with EtOAc (2×). The organics were dried (MgSO₄),filtered, and concentrated. The crude was purified using RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.05% TFA, at flow rate of 30 mL/min) to give 1.4 mg (3.6%)of the desired product as the TFA salt. The product was isolated as asingle diastereomer. LCMS calculated for C₂₃H₃₁ClN₇O₂(M+H)⁺: m/z=472.2;Found: 472.2.

Example 113.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-3,3,3-trifluoropropan-1-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (20 mg, 0.04 mmol, chrial intermediate from peak 1 ofExample 1, step 7) and triethylamine (19 μL, 0.13 mmol) in acetonitrile(0.6 mL) was added 2-bromo-3,3,3-trifluoropropan-1-ol (from SynquestLabs, 9.2 mg, 0.048 mmol). N,N-dimethylformamide (0.3 mL) was added,which created a clear solution that was stirred at 70° C. overnight. Themixture was diluted water and purified using RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 30 mL/min) to give 6.6 mg (30%) ofthe desired product. The product was isolated as a mixture ofdiastereomers. LCMS calculated for C₂₂H₂₇ClF₃N₆O₂ (M+H)⁺: m/z=499.2;Found: 499.1.

Example 115.(2R)-3-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1,1,1-trifluoropropan-2-ol

A mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (20 mg, 0.044 mmol, chrial intermediate from peak 1 ofExample 1, Step 7), (2R)-2-(trifluoromethyl)oxirane (9.4 μL, 0.11 mmol),and triethylamine (18 μL, 0.13 mmol) in ethanol (0.3 mL) was heated in amicrowave at 120° C. for 25 min. The mixture was diluted with MeOH andpurified by RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give 6.2 mg (28%) of the desired product. The product wasisolated as a single enantiomer. LCMS calculated for C₂₂H₂₇ClF₃N₆O₂(M+H)⁺: m/z=499.2; Found: 499.1.

Example 118.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (15 mg, 0.033 mmol, chrial intermediate from Example 1,Step 7, peak 1), mixture of (2S)-2-hydroxypropanoic acid (4.3 μL, 0.049mmol) (L-lactic acid, 85% aq.) and triethylamine (14 μL, 0.098 mmol) inN,N-dimethylformamide (0.2 mL) was addedN,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uroniumhexafluorophosphate (19 mg, 0.049 mmol). The resulting mixture wasstirred overnight at room temperature. The mixture was diluted with MeOHand purified using RP-HPLC (XBridge C18 column, eluting with a gradientof acetonitrile/water containing 0.1% ammonium hydroxide, at flow rateof 30 mL/min) to give 3.0 mg (20%) of the desired product. The productwas isolated as a single enantiomer. LCMS calculated forC₂₂H₂₈ClN₆O₃(M+H)⁺: m/z=459.2; Found: 459.2.

Example 121.(2R)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-oltrifluoroacetate

This compound was prepared using procedures analogous to those forExample 118 (starting from chiral material from Example 1, Step 7, peak1), with (R)-2-hydroxypropanoic acid instead of (2S)-2-hydroxypropanoicacid (4.3 μL, 0.049 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphateinstead of N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uroniumhexafluorophosphate. The crude was purified using RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.05%TFA, at flow rate of 30 mL/min) to give the desired product as the TFAsalt. The product was isolated as a single enantiomer. LCMS calculatedfor C₂₂H₂₈ClN₆O₃(M+H)⁺: m/z=459.2; Found: 459.2.

Example 139. Enantiomers of1-{1-[5-Chloro-2-ethoxy-4-fluoro-3-(1-isopropylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Step 1. 1-(5-Chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone

This compound was prepared according to the procedure of Example 13 Step3, using 1-(5-chloro-4-fluoro-2-hydroxy-3-iodophenyl)ethanone andiodoethane as the starting materials. LCMS calculated for C₁₀H₁₀ClFI₂(M+H)⁺: m/z=342.9; Found: 342.9.

Step 2. tert-Butyl3-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)azetidine-1-carboxylate

A round-bottom flask equipped with a magnetic stir bar and a rubberseptum was charged with lithium chloride (3.9 g, 91 mmol). The flask washeated at 140° C. for 10 min under high vacuum and backfilled withnitrogen after cooling to room temperature. Zinc (6.0 g, 91 mmol) wasadded and the flask was heated at 140° C. for 10 min under high vacuumand backfilled with nitrogen after cooling to room temperature.Tetrahydrofuran (THF) (38 mL) and 1,2-dibromoethane (233 μL, 2.70 mmol)were added via syringe. The mixture was heated at 60° C. for 10 min andthen cooled to room temperature. Chlorotrimethylsilane (68 μL, 0.54mmol) and iodine (69 mg, 0.27 mmol) in THF (1 mL) were added and theresulting mixture was stirred at 60° C. for 10 min then cooled to roomtemperature. A solution of tert-butyl 3-iodoazetidine-1-carboxylate(12.17 g, 42.99 mmol) in THF (10 mL) was then added and the mixturestirred at 40° C. for 1 h and at room temperature for 1 h. Another flaskcharged with 1-(5-chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone (13.0g, 38.0 mmol), palladium acetate (170 mg, 0.76 mmol),2′-(dicyclohexylphosphino)-N,N,N′,N′-tetramethylbiphenyl-2,6-diamine(660 mg, 1.5 mmol), and toluene (35 mL) was evacuated under high vacuumand backfilled with nitrogen. The mixture was cooled to 0° C. and thezinc reagent made above was added slowly via syringe. After addition,the reaction was heated to 50° C. overnight. The reaction solution waspartitioned between EtOAc and sat. NH₄Cl solution. The layers wereseparated and the aqueous extracted further with EtOAc (2×). Thecombined organics were washed with water, brine, then dried over MgSO4,and concentrated. The crude mixture was purified on silica gel column togive the desired product as an orange oil (6.3 g, 45%). LCMS calculatedfor C₁₈H₂₃ClFNO₄Na (M+Na)⁺: m/z=394.1; Found: 394.1.

Step 3. tert-Butyl3-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]azetidine-1-carboxylate

This compound was prepared according to the procedure of Example 13 Step5, using tert-butyl3-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)azetidine-1-carboxylate andsodium tetrahydroborate as the starting materials. LCMS calculated forC₁₈H₂₅ClFNO₄Na (M+Na)⁺: m/z=396.1; Found: 396.1.

Step 4. tert-Butyl3-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-fluorophenyl]azetidine-1-carboxylate

This compound was prepared according to the procedure of Example 13 step6, using tert-butyl3-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]azetidine-1-carboxylate(racemic) and cyanuric chloride as the starting materials.

Step 5. tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidine-1-carboxylate

To a mixture of 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (1.10 g,7.37 mmol), cesium carbonate (3.2 g, 10 mmol) and potassium iodide (111mg, 0.670 mmol) in DMF (20 mL) was added tert-butyl3-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-fluorophenyl]azetidine-1-carboxylate(2.63 g, 6.70 mmol) and the mixture was stirred at 90° C. for 3 h. Thesolvent was removed in vacuo. The residue was diluted with ethyl acetateand water. Aqueous layer was extracted with ethyl acetate twice. Thecombined organic layers were washed with water, brine, dried overNa₂SO₄, filtered and concentrated. The residue was purified on silicagel column (eluting with 100% ethyl acetate) to give the desired productas a foam (2.15 g, 63%). LCMS calculated for C₂₄H₃₁ClFN₆O₃(M+H)⁺:m/z=505.2; Found: 505.2.

Step 6.1-[1-(3-Azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

To a solution of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidine-1-carboxylate(275 mg, 0.544 mmol) in dichloromethane (2.4 mL) was added 4.0 Mhydrogen chloride in dioxane (1.1 mL, 4.4 mmol). The reaction solutionwas stirred at room temperature for 6 h. The solvent was removed underreduced pressure to give the desired product as a white solid (250 mg,96%). LCMS calculated for C₁₉H₂₃ClFN₆O (M+H)⁺: m/z=405.2; Found: 405.1.

Step 7.1-{-[5-Chloro-2-ethoxy-4-fluoro-3-(1-isopropylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (49 mg, 0.10 mmol), acetone (8.28 μL, 0.113 mmol), andtriethylamine (44.3 μL, 0.318 mmol) in dichloromethane (0.67 mL) wasadded sodium triacetoxyborohydride resin (89 mg, 0.20 mmol). Theresulting mixture was stirred overnight at room temperature. The mixturewas filtered and concentrated and then purified by preparative LCMS(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.05% TFA, at flow rate of 60 mL/min) to give the racemicproduct. LCMS: found m/z=447.2 (M+H)⁺. The racemic mixture was separatedby chiral HPLC (column IA, eluting with 5% ethanol/95% hexanes, at flowrate 18 mL/min) to give two peaks (isomer 1: 9.5 mg, 21%; isomer 2: 9.2mg, 20%).

Isomer 1 (first to elute, retention time: 4.4 min): 1H NMR (400 MHz,DMSO-d₆): δ□ 8.10 (s, 1H), 7.45 (d, 1H), 6.21 (m, 1H), 3.70 (m, 5H),2.91 (m, 2H), 2.53 (s, 3H), 2.17 (m, 1H), 1.66 (d, 3H), 1.31 (t, 3H),0.81 (m, 6H) ppm; LCMS calculated for C₂₂H₂₉ClFN₆O (M+H)⁺: m/z=447.2;Found: 447.2.

Isomer 2 (second to elute, retention time: 19.5 min): LCMS calculatedfor C₂₂H₂₉ClFN₆O (M+H)⁺: m/z=447.2; Found: 447.2.

Example 140.1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidin-1-yl)-2-methylpropan-2-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (20 mg, 0.042 mmol, racemic intermediate from Example139, Step 6) and triethylamine (18 μL, 0.12 mmol) in ethanol (1 mL) wasadded oxirane, 2,2-dimethyl-(6.98 μL, 0.0837 mmol). The resultingmixture was heated at 120° C. in microwave reactor for 45 min. Thereaction was diluted with methanol and purified on RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 30 mL/min) to give the desiredproduct as white a solid (3.4 mg, 17%). The product was isolated as aracemic mixture. LCMS calculated for C₂₃H₃₁ClFN₆O₂ (M+H)⁺: m/z=477.2;Found: 477.3.

Example 141.1-(1-{5-Chloro-2-ethoxy-4-fluoro-3-[1-(2,2,2-trifluoroethyl)azetidin-3-yl]phenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (19 mg, 0.040 mmol, racemic intermediate from Example139, Step 6) and triethylamine (20 μL, 0.14 mmol) in dichloromethane(0.5 mL) was added 2,2,2-trifluoroethyl trifluoromethanesulfonate (11mg, 0.048 mmol). The resulting mixture was stirred overnight at roomtemperature. The solvents were evaporated under reduced pressure and thecrude mixture purified on RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 30 mL/min) to give the desired product (3.8 mg, 19%). Theproduct was isolated as a racemic mixture. LCMS calculated forC₂₁H₂₄ClF₄N₆O (M+H)⁺: m/z=487.2; Found: 487.1.

Example 149.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidin-1-yl)propan-2-ol

Step 1. Enantiomers of tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidine-1-carboxylate

The racemic mixture was separated by chiral HPLC (column IA, elutingwith 5% ethanol/95% hexanes, flow rate 18 mL/min) to give two peaks;Isomer 1 (first to elute): Retention time: 16.8 min; LCMS calculated forC₂₄H₃₁ClFN₆O₃(M+H)⁺: m/z=505.2; Found: 505.2; Isomer 2 (second toelute): Retention time: 19.5 min; LCMS calculated for C₂₄H₃₁ClFN₆O₃(M+H)⁺: m/z=505.2; Found: 505.2.

Step 21-[1-(3-Azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

This compound was prepared using procedures analogous to those forExample 139 step 6 with tert-butyl3-{3-[(1S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidine-1-carboxylate(first peak from chiral separation) as starting material. LCMScalculated for C₁₉H₂₃ClFN₆O (M+H)⁺: m/z=405.2; Found: 405.1.

Step 3.(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidin-1-yl)propan-2-ol

To a mixture of1-[1-(3-azetidin-3-yl-5-chloro-2-ethoxy-4-fluorophenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (46 mg, 0.11 mmol) (from isomer 1) and triethylamine (50μL, 0.4 mmol) in isopropyl alcohol (0.3 mL) was added(S)-(−)-methyloxirane (16 μL, 0.23 mmol). The resulting mixture wasstirred at 90° C. for 3 h. After cooling, the mixture was diluted withacetonitrile and purified by RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 30 mL/min) to give the desired product (12 mg, 23%). Theproduct was isolated as a single diastereomer. ¹H NMR (400 MHz,DMSO-d₆): δ 8.05 (s, 1H), 7.38 (d, 1H), 6.15 (m, 1H), 4.26 (d, 1H),3.76-3.60 (m, 6H), 2.99 (m, 2H), 2.48 (s, 3H), 2.22 (m, 2H), 1.62 (d,3H), 1.25 (t, 3H), 0.93 (d, 3H) ppm; LCMS calculated forC₂₂H₂₉ClFN₆O₂(M+H)⁺: m/z=463.2; Found: 463.2.

Example 156.(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propan-1-ol

Step 1. Methyl (2S)-2-bromopropanoate

DMF (28 μL, 0.36 mmol) was added to a mixture of (2S)-2-bromopropanoicacid (0.552 g, 3.61 mmol) and oxalyl chloride (0.61 mL, 7.2 mmol) indichloromethane (4.6 mL) at 0° C. The reaction mixture was stirred atroom temperature overnight. The solvent was removed in vacuo. Theresidue was dissolved in dichloromethane and treated with methanol (1.5mL, 36 mmol) and pyridine (0.44 mL, 5.4 mmol). The reaction solution wasstirred at room temperature for 2 h. The reaction solution was quenchedwith saturated sodium bicarbonate solution and washed with brine, driedover Na₂SO₄, filtered and concentrated to give the desired product (0.51g, 85%).

Step 2. Methyl(2R)-2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propanoate

To a solution of1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminehydrochloride (20.1 mg, 0.0475 mmol, chiral intermediate from Example 1,Step 7) in acetonitrile (1 mL) was added triethylamine (23 μL, 0.17mmol) and methyl (2S)-2-bromopropanoate (9.5 mg, 0.057 mmol). Thereaction solution was stirred at room temperature for 4 h. The solventwas removed to give the desired product (6.2 mg, 28%). LCMS calculatedfor C₂₃H₃₀ClN₆O₃(M+H)⁺: m/z=473.2; Found: 473.3.

Step 3.(2R)-2-(3-{3-[(1S)-1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propan-1-ol

A solution of methyl(2R)-2-(3-{3-[(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propanoate(6.2 mg, 0.013 mmol) in dichloromethane (0.5 mL) was treated with 1.0 Mdiisobutylaluminum hydride in toluene (0.1 mL, 0.1 mmol) at 0° C. for 3h. The reaction was quenched with methanol and purified with preparativeRP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product (0.8 mg, 14%). The product wasisolated as a single diastereomer. LCMS calculated forC₂₂H₃₀ClN₆O₂(M+H)⁺: m/z=445.2; Found: 445.1.

Example 158.1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-2-methylpropan-2-ol

This compound was prepared using procedures analogous to t Example 140with1-[1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminehydrochloride (chiral intermediate from Example 1, Step 7) and oxirane,2,2-dimethyl—as starting materials. The product was isolated as singleenatiomer. LCMS calculated for C₂₃H₃₂ClN₆O₂(M+H)⁺: m/z=459.2; Found:459.1 ¹H NMR (300 MHz, DMSO-d₆): δ 8.04 (s, 1H), 7.23 (bs, 2H), 7.16 (s,1H), 6.14 (m, 1H), 3.96 (s, 1H), 3.85 (m, 3H), 3.45 (s, 3H), 2.94 (m,1H), 2.80 (m, 1H), 2.49 (s, 3H), 2.14 (s, 2H), 2.00 (s, 3H), 1.63 (d,3H), 0.98 (s, 6H) ppm.

Example 159.(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-N,N-dimethylpropanamide

Step 1.(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propanoicacid

To a solution of methyl(2R)-2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propanoate(chiral intermediate from example 156 step 2) (13 mg, 0.027 mmol) inacetonitrile (0.6 mL) and water (0.2 mL) was added lithium hydroxide(2.4 mg, 0.10 mmol). The reaction mixture was stirred at roomtemperature overnight. The reaction solution was diluted with ethylacetate and 1 M HCl solution. The organic layer was separated and driedover Na₂SO₄, filtered and concentrated to give the desired product (10.2mg, 83%). LCMS calculated for C₂₂H₂₈ClN₆O₃(M+H)⁺: m/z=459.2; Found:459.1.

Step 2.(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-N,N-dimethylpropanamide

To a solution of(2R)-2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propanoicacid (4 mg, 0.009 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(4 mg, 0.009 mmol) in DMF (0.3 mL) at room temperature was addedtriethylamine (4 μL, 0.03 mmol) and dimethylamine hydrochloride (0.9 mg,0.01 mmol). The reaction mixture was stirred for 1 h, then diluted withmethanol and purified by preparative RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product (2.7mg, 63%). The product was isolated as a single diastereomer. LCMScalculated for C₂₄H₃₃ClN₇O₂(M+H)⁺: m/z=486.2; Found: 486.1. ¹H NMR (300MHz, DMSO-d₆): δ□ 8.09 (s, 1H), 7.23 (s, 1H), 6.18 (m, 1H), 3.78 (m,3H), 3.50 (s, 3H), 3.01 (s, 3H), 3.0-2.9 (m, 3H), 2.77 (s, 3H), 2.54 (s,3H), 2.06 (s, 3H), 1.67 (d, 3H), 0.98 (d, 3H) ppm.

Example 161.[1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)cyclobutyl]acetonitrile

To a solution of1-[(1-(3-azetidin-3-yl-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (10 mg, 0.022 mmol, chiral intermediate from Example 1,Step 7) in acetonitrile (0.1 mL) was added cyclobutylideneacetonitrile(4.1 mg, 0.044 mmol), followed by 1,8-diazabicyclo[5.4.0]undec-7-ene (13μL, 0.087 mmol). The resulting mixture was stirred at room temperatureovernight. The reaction mixture was diluted with acetonitrile andpurified by preparative RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 30 mL/min) to give the desired product (4.3 mg, 41%). Theproduct was isolated as a single enantiomer. LCMS calculated forC₂₅H₃₁ClN₇O (M+H)⁺: m/z=480.2; Found: 480.0.

Example 163.1-{1-[5-Chloro-2-methoxy-4-methyl-3-(1-methylpiperidin-4-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Step 1. tert-Butyl4-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)piperidine-1-carboxylate

This compound was prepared using procedures analogous to those forExample 139 step 2 with1-(5-chloro-3-iodo-2-methoxy-4-methylphenyl)ethanone and tert-butyl4-iodopiperidine-1-carboxylate as starting materials. LCMS calculatedfor C₂₀H₂₈ClNO₄Na (M+Na)⁺: m/z=404.1; Found: 404.1.

Step 2. tert-Butyl4-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]piperidine-1-carboxylate

This compound was prepared according to the procedure of Example 13 step5, using of tert-butyl4-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)piperidine-1-carboxylateand sodium tetrahydroborate as the starting materials. LCMS calculatedfor C₂₀H₃₀ClNO₄Na (M+Na)⁺: m/z=406.1; Found: 406.1.

Step 3. tert-Butyl4-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]piperidine-1-carboxylate

This compound was prepared according to the procedure of Example 13 step6, using tert-butyl4-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]piperidine-1-carboxylate(racemic) and cyanuric chloride as the starting materials. ¹H NMR (400MHz, CDCl₃): δ 7.44 (s, 1H), 5.46 (m, 1H), 4.23 (bs, 2H), 3.73 (s, 3H),3.29 (bs, 1H), 2.78 (bs, 2H), 2.40 (s, 3H), 2.27-2.09 (m, 2H), 1.78 (d,3H), 1.63 (m, 2H), 1.43 (s, 9H) ppm.

Step 4. tert-Butyl4-{3-[1-(4-amino-3-methyl-H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}piperidine-1-carboxylate

This compound was prepared according to the procedure of Example 139step 5, using of tert-butyl4-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]piperidine-1-carboxylateand 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine as the startingmaterials. LCMS calculated for C₂₆H₃₆ClN₆O₃(M+H)⁺: m/z=515.3; Found:515.2.

Step 5.1-[1-(5-Chloro-2-methoxy-4-methyl-3-piperidin-4-ylphenyl)ethyl]-3-methyl-H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride

This compound was prepared according to the procedure of Example 139step 6, using of tert-butyl4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}piperidine-1-carboxylateas the starting material. LCMS calculated for C₂₁H₂₈ClN₆O (M+H)⁺:m/z=415.2; Found: 415.2.

Step 6.1-{-[5-Chloro-2-methoxy-4-methyl-3-(1-methylpiperidin-4-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

This compound was prepared according to the procedure of Example 139step 7, using of1-[1-(5-chloro-2-methoxy-4-methyl-3-piperidin-4-ylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride and formaldehyde as the starting materials. The productwas isolated as a racemic mixture. LCMS calculated for C₂₂H₃₀ClN₆O(M+H)⁺: m/z=429.2; Found: 429.1.

Example 164.1-(4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}piperidin-1-yl)-2-methylpropan-2-ol

This compound was prepared using procedures analogous to those forExample 140 with1-[1-(5-chloro-2-methoxy-4-methyl-3-piperidin-4-ylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-aminedihydrochloride (racemic intermediate from Example 163, Step 5) andoxirane, 2,2-dimethyl- as starting materials. The product was isolatedas a racemic mixture. LCMS calculated for C₂₅H₃₆ClN₆O₂(M+H)⁺: m/z=487.3;Found: 487.3. ¹H NMR (300 MHz, DMSO-d₆): δ 8.05 (s, 1H), 7.24 (bs, 2H),7.22 (s, 1H), 6.16 (m, 1H), 4.01 (bs, 1H), 3.67 (s, 3H), 2.97 (m, 3H),2.49 (s, 3H), 2.32 (s, 3H), 2.15-2.04 (m, 6H), 1.63 (d, 3H), 1.40 (m,2H), 1.03 (s, 6H) ppm.

Example 166.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}cyclobutanoltrifluoroacetate

Step 1. 1-(5-Chloro-2-methoxy-4-methyl-3-vinylphenyl)ethanone

A mixture of 1-(5-chloro-3-iodo-2-methoxy-4-methylphenyl)ethanone (1.0g, 3.2 mmol, from Example 1, Step 2),4,4,5,5-tetramethyl-2-vinyl-1,3,2-dioxaborolane (0.66 mL, 3.9 mmol),[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (0.26 g, 0.32 mmol) and potassium carbonate(1.3 g, 9.4 mmol) in 1,4-dioxane (10 mL) and water (5 mL) was degassedwith N₂ and heated at 80° C. overnight. After cooled to roomtemperature, the reaction mixture was diluted with water and ethylacetate. The organic layer was washed with brine, dried over MgSO₄,concentrated and purified on a silica gel column (eluting with 0 to 10%EtOAc in hexanes) to give the desired product (0.60 g, 82%). LCMScalculated for C₁₂H₁₄ClO₂ (M+H)⁺: m/z=225.1; Found: 225.1

Step 2. 3-(3-Acetyl-5-chloro-2-methoxy-6-methylphenyl)cyclobutanone

To a solution of 1-(5-chloro-2-methoxy-4-methyl-3-vinylphenyl)ethanone(530 mg, 2.4 mmol) in ether (10 mL) was added zinc-copper couple (1.8 g,14 mmol). The reaction mixture was heated at 40° C. and a solution oftrichloroacetyl chloride (1.4 mL, 13 mmol) and phosphoryl chloride (1.2mL, 13 mmol) in 1,2-dimethoxyethane (3 mL) was added slowly over 2 h.After addition, the reaction mixture was stirred under reflux overnight.The reaction was quenched with saturated NaHCO₃ solution and dilutedwith ether. The organic layer was washed with brine, dried over Na₂SO₄,filtered and concentrated. The residue and zinc (0.31 g, 4.7 mmol) inacetic acid (10 mL) was stirred at room temperature for 2 h and thenreflux overnight. Another portion of zinc was added and reflux foranother 4 h. The mixture was diluted with water and extracted withether. The organic phase was washed successively with a saturated NaHCO₃solution, water and brine, then dried over MgSO₄ and concentrated. Thecrude material was purified with flash chromatography (eluting with 0 to30% ethyl acetate in hexanes) to give the desired product (0.17 g, 27%).LCMS calculated for C₁₄H₁₆ClO₃ (M+H)⁺: m/z=267.1; Found: 267.0.

Step 3.3-[3-Chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]cyclobutanol

This compound was prepared according to the procedure of Example 13 step5, using of 3-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)cyclobutanoneand sodium tetrahydroborate as the starting materials. LCMS calculatedfor C₁₄H₁₉ClO₃Na (M+Na)⁺: m/z=293.1; Found: 293.1.

Step 4.3-[3-Chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]cyclobutanol

To a solution of3-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]cyclobutanol(170 mg, 0.628 mmol) in dimethyl sulfoxide (1 mL) was added cyanuricchloride (64 mg, 0.34 mmol). After stirred overnight, the reactionmixture was diluted with ether and water. The aqueous layer wasextracted with ethyl acetate once. The combined organic extracts werewashed with brine, dried over Na₂SO₄, filtered and concentrated. Thecrude was purified with silica gel column to give the desired product(39.6 mg, 22%). LCMS calculated for C₁₄H₁₈ClO₂ (M-Cl)⁺: m/z=253.1;Found: 253.2.

Step 5.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}cyclobutanoltrifluoroacetate

This compound was prepared according to the procedure of Example 139step 5, using of3-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]cyclobutanol and3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine as the starting materials.The product was isolated as a racemic mixture. LCMS calculated forC₂₀H₂₅ClN₅O₂(M+H)⁺: m/z=402.2; Found: 402.2.

Example 167.5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

Step 1. 1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone

To a stirred solution of 1-(5-chloro-2-methoxy-4-methylphenyl)ethanone(5.00 g, 25.2 mmol, from Oakwood) in acetic acid (100 mL) was addedN-bromosuccinimide (4.93 g, 27.7 mmol) and the resulting mixture heatedat 100° C. for 18 hours. After cooling to ambient temperature, thereaction mixture was concentrated in vacuo, then neutralized with sat.sodium bicarbonate, filtered off insoluble succinimide. The filtrate wasextracted with EtOAc. The combined organic layers were washed withbrine, dried over sodium sulfate, and then concentrated to dryness underreduced pressure. The residue was purified on silica gel, eluting with 0to 50% EtOAc in hexanes, to give the desired products (2.66 g, 38%).LCMS calculated for C₁₀H₁₁BrClO₂ (M+H)⁺: m/z=277.0; found: 277.0. ¹H NMR(DMSO-d₆, 300 MHz): δ 7.70 (1H, s), 3.77 (3H, s), 2.57 (3H, s), 2.50(3H, s) ppm.

Step 2. 1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl)ethanol

Sodium tetrahydroborate (0.31 g, 8.1 mmol) was added to a mixture of1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone (1.5 g, 5.4 mmol)in methanol (25 mL) at 0° C. and the resultant reaction mixture wasstirred at room temperature for 1 hour. The solvent was removed and theresulting residue was diluted with ethyl acetate, washed with sat.NaHCO₃, water, brine, then dried over Na₂SO₄, filtered and concentrated.The crude product was purified by silica gel chromatography, elutingwith 0 to 40% EtOAc in hexanes, to give the desired product (0.30 g,90%).

Step 3. 3-Bromo-1-chloro-5-(1-chloroethyl)-4-methoxy-2-methylbenzene

A mixture of cyanuric chloride (1.7 g, 9.2 mmol) andN,N-dimethylformamide (710 μL, 9.2 mmol) was stirred at room temperaturefor 10 minutes and then a solution of1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanol (from Example 16,Step 1) (1.72 g, 6.15 mmol) in methylene chloride (34 mL) was added andthe reaction was stirred at room temperature overnight. The mixture wasdiluted with methylene chloride, washed with sat. NaHCO₃, water, brine,dried over Na₂SO₄, filtered and concentrated. The crude product waspurified by silica gel chromatography, eluting with 0 to 10% EtOAc inhexanes, to give the desired product (1.01 g, 60%).

Step 4.1-[1-(3-Bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A mixture of3-bromo-1-chloro-5-(1-chloroethyl)-4-methoxy-2-methylbenzene (150 mg,0.503 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (110 mg, 0.76mmol, ACES Phanna Product List, item #47024), potassium iodide (9.0 mg,0.05 mmol) and cesium carbonate (330 mg, 1.0 mmol) inN,N-dimethylformamide (4 mL) and was stirred at 140° C. for 1 h. Themixture was diluted with methylene chloride, washed with sat. NaHCO₃,water, brine, dried over Na₂SO₄, filtered and concentrated. The crudeproduct was purified by silica gel chromatography, eluting with 0 to 70%EtOAc in CH₂Cl₂, to give the desired product (103 mg, 50%). LCMScalculated for C₁₆H₁₈BrClN₅O (M+H)⁺: m/z=410.0; Found: 410.2. Theracemic products were applied on a Phenomenex Lux-Cellulose 1 column(21.1×250 mm, 5 micron particle size), eluting with 5% ethanol inhexanes at a flow rate of 18 mL/min, ˜13 mg/injection, to provide twoenantiomers. Peak 1, retention time: 12.35 min; Peak 2, retention time:14.98 min.

Step 5.5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(25 mg, 0.061 mmol) (first peak from previous step chiral separation),N,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carboxamide(25 mg, 0.091 mmol, from PepTech Corp. Encyclopedia of Amino AcidAnalogs and Boronic Acids, item #BE1622-1), sodium carbonate (13 mg,0.12 mmol) and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complex with dichloromethane (1:1) (9.9 mg, 0.012 mmol) inacetonitrile (0.8 mL)/water (0.3 mL) was degassed with N₂ and thenstirred at 95° C. for 2 h. After cooling to room temperature, themixture was filtered and the filtrate purified on RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.05%trifluoroacetic acid, at flow rate of 30 mL/min) to give the desiredproduct as bis-TFA salt (2.9 mg, 6.7%). The product was isolated as asingle enantiomer. LCMS calculated for C₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2;Found: 480.2. ¹H NMR (500 MHz, DMSO-d₆) δ: 8.78 (2H, br s), 8.48 (1H,m), 8.36 (1H, s), 7.86 (1H, br s), 7.65 (1H, br s), 7.58 (1H, s), 6.33(1H, q, J=7.0 Hz), 3.19 (3H, s), 3.03 (3H, s), 2.97 (3H, s), 2.62 (3H,s), 2.06 (3H, s), 1.81 (3H, d, J=7.0 Hz) ppm.

Example 183.1-[1-(5-Chloro-3-{1-[2-(dimethylamino)ethyl]-1H-pyrazol-4-yl}-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Step 1.1-(2-Chloroethyl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole

A mixture of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole(0.39 g, 2.0 mmol), 1-bromo-2-chloroethane (0.3 mL, 3 mmol) and cesiumcarbonate (1.3 g, 4.0 mmol) in acetonitrile (6 mL) was stirred at 75° C.for 5 h. The mixture was diluted with ethyl acetate, washed with sat.NaHCO₃, water, brine, dried over Na₂SO₄, filtered and concentrated andthe product (0.45 g, 88%) was purified by chromatography eluting withhexanes/EtOAc (max. EtOAc 30%). LCMS calculated for C₁₁H₁₉BClN₂O₂(M+H)⁺:m/z=257.1; Found: 257.0

Step 2.N,N-Dimethyl-2-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl]ethanamine

A mixture of1-(2-chloroethyl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole(0.10 g, 0.39 mmol), sodium iodide (58 mg, 0.39 mmol) and 2.0 Mdimethylamine in THF (1.0 mL, 2.0 mmol) in N,N-dimethylformamide (0.5mL) was stirred at 80° C. overnight. The solvent was removed to providethe desired product which was used in the next step. LCMS calculated forCl₃H₂₅BN₃O₂(M+H)⁺: m/z=266.2; Found: 266.3.

Step 3.1-[1-(5-chloro-3-{1-[2-(dimethylamino)ethyl]-H-pyrazol-4-yl}-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(Peak 1 from Example 167, step 4, 10 mg, 0.024 mmol),N,N-dimethyl-2-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl]-ethanamine(8.6 mg, 0.036 mmol), sodium carbonate (5.2 mg, 0.049 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (4.0 mg, 0.0049 mmol) in acetonitrile (0.5mL)/water (0.1 mL) was vacuumed and the refilled with N₂ and the stirredat 95° C. for 2 h. The crude was purified using RP-HPLC (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 30 mL/min) to give the desiredproduct (3.1 mg, 28%). The product was isolated as a single enantiomer.LCMS calculated for C₂₃H₃₀ClN₈O (M+H)⁺: m/z=469.2; Found: 469.2.

Example 184.2-[(5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridin-2-yl)amino]ethanol

Step 1.1-{1-[5-Chloro-3-(6-fluoropyridin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(Peak 1 from Example 167, step 4, 25.0 mg, 0.06 mmol),2-fluoro-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (20.mg, 0.088 mmol), sodium carbonate (12 mg, 0.12 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (9.5 mg, 0.012 mmol) in acetonitrile (1mL)/water (0.3 mL) was degassed with N₂ and the stirred at 95° C. for 2h. The mixture was diluted with methylene chloride, washed with sat.NaHCO₃, water, brine, dried over Na₂SO₄, filtered and concentrated. Theproduct was purified by chromatography eluting with CH₂Cl₂/MeOH (max.MeOH 5%). LCMS calculated for C₂₁H₂₁ClFN₆O (M+H)⁺: m/z=427; Found:427.2.

Step 2.2-[(5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridin-2-yl)amino]ethanol

A mixture of1-{1-[5-chloro-3-(6-fluoropyridin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(10 mg, 0.023 mmol) and ethanolamine (0.10 mL) in 1-butanol (1 mL) wasstirred at 130° C. for 5 h. The crude was purified using RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 30 mL/min) to givethe desired product (1.6 mg, 15%). The product was isolated as a singleenantiomer. LCMS calculated for C₂₃H₂₇ClN₇O₂ (M+H)⁺: m/z=468.2; Found:468.2.

Example 188.2-(5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)pyridin-2-yloxy)ethanol

Sodium hydride (20 mg, 0.5 mmol) was added to 1,2-ethanediol (0.5 mL, 9mmol) and the mixture was stirred at room temperature for 10 min. Atthis time1-{1-[5-chloro-3-(6-fluoropyridin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(10 mg, 0.023 mmol) was added and then the reaction was stirred at 110°C. overnight. The crude was purified using RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product (1.8mg, 17%). The product was isolated as a single enantiomer. LCMScalculated for C₂₃H₂₆ClN₆O₃(M+H)⁺: m/z=469.2; Found: 469.1.

Example 189.5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-(2,2-difluoroethoxy)-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

Step 1.5-(3-(1-(4-Amino-3-methyl-H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-hydroxy-6-methylphenyl)-N,N-dimethylpicolinamide

1.0 M Boron tribromide in CH₂Cl₂ (250 μL, 0.25 mmol) was added to amixture of5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide(Example 167, step 5, (first peak) 60 mg, 0.13 mmol) in methylenechloride (1.2 mL) at −78° C. and then the reaction was warmed to roomtemperature. At this time conc. HCl (0.1 mL) was added and the mixturewas stirred for 4 h. The reaction was quenched by the addition of sat.NaHCO₃. The mixture was then extracted with methylene chloride. Thecombined extracts were washed with brine, dried and concentrated to givethe desired crude product (40 mg, 68%) which was used in the next stepwithout further purification. LCMS calculated for C₂₃H₂₅ClN₇O₂(M+H)⁺:m/z=466.2; Found: 466.2.

Step 2.5-[3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-(2,2-difluoroethoxy)-6-methylphenyl]-NN-dimethylpyridine-2-carboxamide

Diisopropyl azodicarboxylate (13 μL, 0.064 mmol) was added to a mixtureof5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-hydroxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide(15.0 mg, 0.0322 mmol), 2,2-difluoroethanol (7.9 mg, 0.096 mmol, fromAlfa Aesar, item # B22201) and triphenylphosphine (17 mg, 0.064 mmol) intetrahydrofuran (0.5 mL) at 0° C. and then the reaction was stirred atroom temperature for 24 h. The crude was purified on RP-HPLC (XBridgeC18 column, eluting with a gradient of acetonitrile/water containing0.05% trifluoroacetic acid, at flow rate of 30 mL/min) to give thedesired product as bis-TFA salt (1.6 mg, 6.6%). The product was isolatedas a single enantiomer. LCMS calculated for C₂₅H₂₇ClF₂N₇O₂ (M+H)⁺:m/z=530.2; Found: 530.2.

Example 192.1-[1-(5-Chloro-3-cyclopropyl-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

To a microwave vial was added1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(15 mg, 0.037 mmol, from peak 1 from Example 167, step 4), potassiumcyclopropyltrifluoroborate (8 mg, 0.06 mmol, from Frontier Scientific,item # C10298), potassium phosphate (23 mg, 0.11 mmol), andtetrakis(triphenylphosphine)palladium (4.2 mg, 0.0036 mmol) and thentoluene (0.3 mL)/water (0.1 mL). The vial was sealed and degassed withN₂ three times. The reaction was heat at 110° C. for 20 h. The crude waspurified using RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product (1.1 mg, 8%). The product wasisolated as a single enantiomer. LCMS calculated for C₁₉H₂₃ClN₅O (M+H)⁺:m/z=372.2; Found: 372.2.

Example 195.5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

Step 1. 1-(3-Bromo-5-chloro-2-ethoxy-4-methylphenyl)ethanone

Into a round bottom flask was placed1-(3-bromo-5-chloro-2-hydroxy-4-methylphenyl)ethanone (6.0 g, 23 mmol)in anhydrous DMF (22.8 mL). Potassium carbonate (6.3 g, 46 mmol) wasthen added followed by iodoethane (2.73 mL, 34.2 mmol). The resultingsuspension was stirred at 60° C. for 2 h. The mixture was poured into100 mL water and extracted with 200 mL of ethyl ether. The organiclayers were separated, combined and washed with water and saturated NaClsolution, dried over anhydrous sodium sulfate, filtered, andconcentrated to 6.0 g of tan oil. LCMS calculated for C₁₁H₁₃BrClO₂(M+H)⁺: m/z=293.0; Found: 293.0.

Step 2. 1-(3-Bromo-5-chloro-2-ethoxy-4-methylphenyl)ethanol

Sodium tetrahydroborate (0.31 g, 8.1 mmol) was added to a mixture of1-(3-bromo-5-chloro-2-ethoxy-4-methylphenyl)ethanone (1.5 g, 5.4 mmol)in methanol (25 mL) at 0° C. and the resultant reaction mixture wasstirred at room temperature for 1 hour. The solvent was removed and theresulting residue was diluted with ethyl acetate, washed with sat.NaHCO₃, water, brine, then dried over Na₂SO₄, filtered and concentrated.The crude product was purified by silica gel chromatography, elutingwith 0 to 30% EtOAc in hexanes (0.30 g, 90%).

Step 3. 3-Bromo-1-chloro-5-(1-chloroethyl)-4-ethoxy-2-methylbenzene

A mixture of cyanuric chloride (1.7 g, 9.2 mmol) andN,N-dimethylformamide (710 μL, 9.2 mmol) was stirred at room temperaturefor 10 minutes and then a solution of1-(3-bromo-5-chloro-2-ethoxy-4-methylphenyl)ethanol (1.72 g, 6.15 mmol)in methylene chloride (34 mL) was added and the reaction was stirred atroom temperature overnight. The mixture was diluted with methylenechloride, washed with sat. NaHCO₃, water, brine, dried over Na₂SO₄,filtered and concentrated. The crude product was purified by silica gelchromatography, eluting with 0 to 10% EtOAc in hexanes (1.01 g, 60%).

Step 4.1-(1-(3-Bromo-5-chloro-2-ethoxy-4-methylphenyl)ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A mixture of 3-bromo-1-chloro-5-(1-chloroethyl)-4-ethoxy-2-methylbenzene(150 mg, 0.50 mmol), 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (110mg, 0.76 mmol), potassium iodide (9 mg, 0.05 mmol) and cesium carbonate(330 mg, 1.0 mmol) in N,N-dimethylformamide (4 mL) was stirred at 140°C. for 1 h. The mixture was diluted with methylene chloride, washed withsat. NaHCO₃, water, brine, dried over Na₂SO₄, filtered and concentrated.The crude product was purified by silica gel chromatography, elutingwith 0 to 70% EtOAc in CH₂Cl₂ (103 mg, 50%). LCMS calculated forC₁₇H₂₀BrClN₅O (M+H)⁺: m/z=423.1; Found: 423.0. The racemic products wereapplied on a Phenomenex Lux-Cellulose 1 column (21.1×250 mm, 5 micronparticle size), eluting with 4% ethanol in hexanes at a flow rate of 18mL/min, ˜13 mg/injection, to provide two enantiomers. Peak 1, retentiontime: 8.64 min; Peak 2, retention time: 10.64 min.

Step 5.5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

A mixture of1-[1-(3-bromo-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(25 mg, 0.061 mmol) (first peak from previous step chiral separation),N,N-dimethyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carboxamide(25 mg, 0.09 mmol), sodium carbonate (13 mg, 0.12 mmol) and[1,1′-bis(diphenylphosphino)-ferrocene]dichloropalladium (II), complexwith dichloromethane (1:1) (9.9 mg, 0.012 mmol) in acetonitrile (0.8mL)/water (0.3 mL) was degassed with N₂ and then stirred at 95° C. for 2hours. After cooling to room temperature, the mixture was filtered andthe filtrate purified on RP-HPLC (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.05% trifluoroacetic acid, atflow rate of 30 mL/min) to give the desired product as bis-TFA salt (2.3mg, 5%). The product was isolated as a single enantiomer. LCMScalculated for C₂₅H₂₉ClN₇O₂(M+H)⁺: m/z=494.2; Found: 494.2.

Example 200.4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoro acetate)

Step 1.4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile

A mixture of1-[1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(peak 1 from Example 167, step 4, 322 mg, 0.76 mmol),4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-2-carbonitrile(210 mg, 0.91 mmol, from Combi-Blocks Catalog, item # PN-0143), sodiumcarbonate (130 mg, 1.2 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]-dichloropalladium(II), complexwith dichloromethane (1:1) (99 mg, 0.12 mmol) in acetonitrile (5mL)/water (2 mL) was degassed with N₂ and the reaction was stirred at95° C. for 2 h. The mixture was diluted with methylene chloride, washedwith sat. NaHCO₃, water, brine, dried over Na₂SO₄, filtered andconcentrated. The product (0.28 g, 85%) was purified by chromatographyeluting with CH₂Cl₂/MeOH (max. MeOH 6%). LCMS calculated for C₂₂H₂₁ClN₇O(M+H)⁺: m/z=434.1; Found: 434.1.

Step 2.4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)picolinicacid dihydrochloride

1.0 M Sodium hydroxide (2.9 mL, 2.9 mmol) was added to a mixture of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carbonitrile(0.250 g, 0.576 mmol) in ethanol (4.0 mL) and the resulting mixture washeated at 95° C. for 6 h. At this time, conc. HCl was added to adjustthe pH to ˜3. The solvent was removed and the residue was used in thenext step without further purification. LCMS calculated forC₂₂H₂₂ClN₆O₃(M+H)⁺: m/z=453.1; Found: 453.2.

Step 3.4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamidebis(2,2,2-trifluoroacetate)

2.0 M Dimethylamine in THF (2.0 mL, 4.0 mmol) was added to a solution of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridine-2-carboxylicacid (250 mg, 0.552 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(370 mg, 0.83 mmol) in N,N-dimethylformamide (4 mL) at 0° C. followed byadding triethylamine (0.23 mL, 1.6 mmol). The reaction was stirred for 1h. The crude mixture was purified on RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.05%trifluoroacetic acid, at flow rate of 30 mL/min) to give the desiredproduct as bis-TFA salt. The product was isolated as a singleenantiomer. LCMS calculated for C₂₄H₂₇ClN₇O₂(M+H)⁺: m/z=480.2; Found:480.2. ¹H NMR (DMSO-d₆, 500 MHz) δ 8.67 (br s, 1H), 8.36 (s, 1H), 7.58(s, 1H), 7.41 (m, 2H), 6.32 (q, 2H), 3.20 (s, 3H), 3.00 (s, 3H), 2.94(s, 3H), 2.62 (s, 3H), 2.03 (s, 3H), 1.80 (d, 3H) ppm.

Example 203.2-(4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-methylphenyl)-1H-pyrazol-1-yl)acetamide

Step 1. tert-Butyl[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl]acetate

1.0 M Potassium tert-butoxide in THF (2.4 mL, 2.4 mmol) was added to asolution of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole(0.39 g, 2.0 mmol) in N,N-dimethylformamide (6.0 mL) at 0° C. Thereaction mixture was stirred at room temperature for 5 min. After cooledto 0° C., to the mixture was added t-butyl bromoacetate (0.5 mL, 3mmol). The reaction was stirred at room temperature for 2 h, thendiluted with ethyl acetate, washed with sat. NaHCO₃, water, brine, driedover Na₂SO₄, filtered and concentrated. The product (0.5 g, 81%) waspurified by chromatography eluting with hexanes/EtOAc (max. EtOAc 30%).LCMS calculated for C₁₅H₂₆BN₂O₄(M+H)⁺: m/z=309.2; Found: 309.1.

Step 2. tert-Butyl(4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-1H-pyrazol-1-yl)acetate

A mixture of1-[1-(3-bromo-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(70 mg, 0.16 mmol) (first peak from Example 195, step 4), tert-butyl[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazol-1-yl]acetate(65 mg, 0.21 mmol), sodium carbonate (30. mg, 0.28 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (23 mg, 0.028 mmol) in acetonitrile (3mL)/water (0.7 mL) was degassed with N₂ and then stirred at 95° C. for 2h. The mixture was diluted with methylene chloride, washed with sat.NaHCO₃, water, brine, dried over Na₂SO₄, filtered and concentrated. Theproduct (65 mg, 78%) was purified by chromatography eluting withCH₂Cl₂/MeOH (max. MeOH 5%). LCMS calculated for C₂₆H₃₃ClN₇O₃ (M+H)⁺:m/z=526.2; Found: 526.3.

Step 3.(4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-1H-pyrazol-1-yl)aceticacid bis trifluoroacetate

Trifluoroacetic acid (0.5 mL) was added to a solution of tert-butyl(4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-1H-pyrazol-1-yl)acetate(0.065 g, 0.12 mmol) in methylene chloride (0.5 mL). The reaction wasstirred at room temperature for 4 h. The solvent was removed to providethe crude product which was used in the next step. LCMS calculated forC₂₂H₂₅ClN₇O₃(M+H)⁺: m/z=470.2; Found: 470.1.

Step 4.2-(4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-1H-pyrazol-1-yl)acetamide

Ammonium carbonate (20 mg, 0.21 mmol) was added to a solution of(4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-1H-pyrazol-1-yl)aceticacid bis trifluoroacetate (10 mg, 0.021 mmol) andbenzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate(10 mg, 0.03 mmol) in N,N-dimethylformamide (0.7 mL) at room temperaturefollowed by triethylamine (8.8 μL, 0.064 mmol). The reaction was stirredfor 1 h. The crude was purified using RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product (2.5mg, 25%). The product was isolated as a single enantiomer. LCMScalculated for C₂₂H₂₆ClN₈O₂(M+H)⁺: m/z=469.2; Found: 469.2.

Example 208.6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylnicotinamidebis(trifluoroacetate)

Step 1.1-{1-[5-Chloro-2-ethoxy-4-methyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]ethyl}-3-methyl-H-pyrazolo[3,4-d]pyrimidin-4-amine

1-[1-(3-Bromo-5-chloro-2-ethoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(0.050 g, 0.12 mmol, Peak 1 from Example 195, step 4) was combined in amicrowave vial with potassium acetate (0.035 g, 0.35 mmol) and4,4,5,5,4′,4′,5′,5′-octamethyl-[2,2′]bi[[1,3,2]dioxaborolanyl] (0.060 g,0.24 mmol) in dimethyl sulfoxide (0.44 mL) at room temperature. This wasdegassed with nitrogen and then[1,1′-bis(diphenylphosphino)ferrocene]-dichloropalladium(II), complexwith dichloromethane (1:1) (0.01 g, 0.01 mmol) was added. The reactionwas heated in an oil bath to 105° C. overnight. This was allowed tocool, then taken up in ethyl acetate and washed with water, brine, driedover magnesium sulfate and concentrated. The product (15 mg, 20%) waspurified by chromatography eluting with CH₂Cl₂/MeOH (max. MeOH 10%).LCMS calculated for C₂₃H₃₂BClN₅O₃(M+H)⁺: m/z=472.2; Found: 472.3.

Step 2.6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylnicotinamidebis(trifluoroacetate)

A mixture of1-{1-[5-chloro-2-ethoxy-4-methyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(15 mg, 0.032 mmol), 6-chloro-N,N-dimethylnicotinamide (12 mg, 0.064mmol), sodium carbonate (9.0 mg, 0.085 mmol) and[1,1′-bis(diphenylphosphino)ferrocene]-dichloropalladium(II), complexwith dichloromethane (1:1) (6.9 mg, 0.0085 mmol) in acetonitrile (0.9mL)/water (0.2 mL) was degassed with N₂ and then stirred at 95° C.overnight. The crude was purified using RP-HPLC (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.05%trifluoroacetic acid, at flow rate of 30 mL/min) to give the desiredproduct as TFA salt (2 mg, 9%). The product was isolated as a singleenantiomer. LCMS calculated for C₂₅H₂₉ClN₇O₂ (M+H)⁺: m/z=494.2; Found:494.2.

Example 209.5-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-4-methoxy-2-methyl-3-(1-methyl-1H-pyrazol-4-yl)benzonitrile

Pre-formed catalyst (0.05 mL, from Example 40) was added to a mixture1-{1-[5-chloro-2-methoxy-4-methyl-3-(1-methyl-1H-pyrazol-4-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(7.7 mg, 0.019 mmol), zinc (0.54 mg, 0.0082 mmol) and zinc cyanide (2.2mg, 0.019 mmol) in N,N-dimethylacetamide (0.3 mL). The mixture wasdegassed with nitrogen 3 times. The reaction was heated at 120° C. for1.5 h. The crude was purified using RP-HPLC (XBridge C18 column, elutingwith a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 30 mL/min) to give the desired product (2.1mg, 27%). The product was isolated as a single enantiomer. LCMScalculated for C₂₁H₂₃N₈O (M+H)⁺: m/z=403.2; Found: 403.2.

Experimental procedures and LCMS mass spectral data (MS) for thecompounds below are summarized in Table 1.

TABLE 1

Ex. MS No. Name R² R⁴ R⁵ R³ Salt Proc.¹ [M + H]⁺ 168 1-(1-(5- chloro-2-methoxy-4- methyl-3- (pyrimidin-5- yl)phenyl)eth- yl)-3-methyl- 1H-pyrazolo[3,4- Me Me Cl

2TFA 167 425.1 d]pyrimidin- 4-amine² 169 1-(1-(3-(2- amino- pyrimidin-5-yl)-5-chloro- 2-methoxy-4- methylphen- yl)ethyl)-3- methyl-1H-pyrazolo[3,4- d]pyrimidin- 4-amine² Me Me Cl

167 425.1 ¹Synthesized according to the experimental procedure ofcompound listed; ²Compound isolated as a single enantiomer.

Example 212.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile

Step 1. 1-(5-Chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone

The desired compound was prepared according to the procedure of Example13, step 3 to form a racemic intermediate, using iodoethane instead ofiodomethane as the starting material in 90% yield. ¹H NMR (300 MHz,CDCl₃) δ 7.68 (d, J=8.3 Hz, 1H), 3.94 (q, J=7.0 Hz, 2H), 2.61 (s, 3H),1.48 (t, J=7.0 Hz, 3H). LCMS for C₁₀H₁₀ClFIO₂ (M+H)⁺: m/z=342.9, 344.9;Found: 342.9, 344.8.

Step 2. 4-Acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile

A solution of 1-(5-chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone (7.3g, 21 mmol) in N,N-dimethylformamide (80 mL) was treated with potassiumcyanide (2.1 g, 32 mmol) and stirred at 40° C. for 5 h. The reactionmixture was diluted with ethyl acetate and poured into saturated sodiumbicarbonate solution/water (1:1). The organic layer was separated,washed with saturated sodium bicarbonate solution, dried with magnesiumsulfate, filtered, and concentrated to give a crude brown oil. The crudematerial was purified by flash column chromatography using ethyl acetatein hexanes (0%-30%) to give the desired product (6.1 g, 81%) as a yellowsolid. ¹H NMR (400 MHz, CDCl₃) δ 7.57 (s, 1H), 3.93 (q, J=7.0 Hz, 2H),2.61 (s, 3H), 1.47 (t, J=7.0 Hz, 3H). LCMS for Cl₁H₁₀ClINO₂ (M+H)⁺:m/z=349.9; Found: 349.9.

Step 3. tert-Butyl3-(3-acetyl-5-chloro-6-cyano-2-ethoxyphenyl)azetidine-1-carboxylate

Zinc (4.60 g, 70.3 mmol) and oven dried Celite (870 mg) was added to aflask and the flask was heated with a heat gun while under high-vac for5 min and then back-filled with nitrogen. N,N-Dimethylacetamide (57 mL)was added, followed by 1,2-dibromoethane (430 μL, 5.0 mmol) and themixture was heated at 70° C. for 10 min and then cooled to roomtemperature. The reaction mixture was treated with chlorotrimethylsilane(630 μL, 5.0 mmol) dropwise and stirred at room temperature for 1 h. Thereaction mixture was treated with a solution of tert-butyl3-iodoazetidine-1-carboxylate (18 g, 62 mmol) in N,N-dimethylacetamide(28 mL) dropwise (internal temperature was kept below 40° C. with awater bath) and heated at 40° C. for 2 h. The zinc-iodo reagent(transferred via canula) was filtered through a plastic filter (that wasappropriately sealed to avoid atmospheric exposure) directly into aclean, dry flask that was flushed with nitrogen. The reaction mixturewas treated with tris(dibenzylideneacetone)dipalladium(0) (720 mg, 0.79mmol) and tri-(2-furyl)phosphine (370 mg, 1.6 mmol) and degassed withnitrogen for a few minutes. The reaction mixture was treated with asolution of 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile (14 g, 41mmol) in N,N-dimethylacetamide (130 mL) (degassed with nitrogen) quicklyand heated at 70° C. for 2 h. The reaction mixture was poured intosaturated ammonium chloride solution and extracted with ethyl acetate(3×300 mL). The combined organic extracts were washed with water (4×500mL) and brine (1×500 mL), dried with magnesium sulfate, filtered, andconcentrated to a crude dark oil. The crude material was purified byflash column chromatography using ethyl acetate in hexanes (5%-45%) togive the desired product (14 g, 88%). ¹H NMR (300 MHz, CDCl₃) δ 7.46 (s,1H), 4.42-4.20 (m, 5H), 3.80 (q, J=7.0 Hz, 2H), 2.59 (s, 3H), 1.44 (s,9H), 1.37 (t, J=7.0 Hz, 3H). LCMS for C₁₅H₁₆ClN₂O₄ ([M-(t-Bu)+H]+H)⁺:m/z=323.1; Found: 323.0.

Step 4. tert-Butyl3-[3-chloro-2-cyano-6-ethoxy-5-(1-hydroxyethyl)phenyl]azetidine-1-carboxylate

A solution of(3aS)-1-methyl-3,3-diphenyltetrahydro-3H-pyrrolo[1,2-c][1,3,2]oxazaborole(9.7 g, 35 mmol) in tetrahydrofuran (100 mL) was treated with 1.0 Mborane-THF complex in tetrahyrofuran (42 mL, 42 mmol) and stirred at 20°C. for 15 min. The reaction mixture was cooled to −30° C. and treatedwith a solution of tert-butyl3-(3-acetyl-5-chloro-6-cyano-2-ethoxyphenyl)azetidine-1-carboxylate (13g, 35 mmol) in tetrahydrofuran (110 mL) slowly. The flask containing thestarting material ketone was rinsed with additional tetrahydrofuran (20mL) and added to the reaction mixture. The reaction mixture was warmedto 0° C. over a period of 30 min and stirred at 0° C. for 15 min. Thereaction mixture was quenched with water at 0° C., poured into saturatedsodium bicarbonate solution, and extracted with ethyl acetate. Theaqueous layer was separated and extracted with ethyl acetate. Thecombined organic layers were washed with water and brine, dried withmagnesium sulfate, filtered, and concentrated to a crude dark oil. Thecrude material was purified by flash column chromatography using ethylacetate in hexanes (0%-70%) to give the desired product (10.4 g, 78%) asa yellow foam as a 98:2 mixture of enantiomers (Retention times=7.73 minand 9.41 min; ChiralPak AD-H column, 4.6×150 mm, 5 micron particle size,eluting with 5% ethanol in hexanes at 1 ml/min). ¹H NMR (300 MHz, CDCl₃)δ 7.56 (s, 1H), 5.15-5.07 (m, 1H), 4.41-4.17 (m, 5H), 3.74 (q, J=7.0 Hz,2H), 2.12 (d, J=3.7 Hz, 1H), 1.49-1.37 (m, 15H). LCMS for C₁₅H₁₈ClN₂O₄([M-(t-Bu)+H]+H)⁺: m/z=325.1; Found: 325.1.

Step 5. tert-Butyl3-{3-[1-(4-amino-3-methyl-H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidine-1-carboxylate

A solution of tert-butyl3-[3-chloro-2-cyano-6-ethoxy-5-(1-hydroxyethyl)phenyl]azetidine-1-carboxylate(98:2 mixture of enantiomers from step 4) (10 g, 27 mmol) in methylenechloride (260 mL) at 0° C. was treated with triethylamine (11 mL, 82mmol) followed by methanesulphonic anhydride (7.1 g, 41 mmol) andstirred at 0° C. for 15 min. The reaction mixture was diluted withdichloromethane and washed with water and brine, dried with magnesiumsulfate, filtered, and concentrated to give the crude mesylate that wasused without further purification. A solution of the crude mesylateintermediate in N,N-dimethylformamide (140 mL) was treated with cesiumcarbonate (13 g, 41 mmol) and3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (4.7 g, 31 mmol) and heatedat 60° C. for 1 h. The reaction mixture was diluted with water andextracted with ethyl acetate (3×250 mL). The combined organic layerswere washed with water and brine, dried with magnesium sulfate,filtered, and concentrated to a crude oil. The crude material waspurified by flash column chromatography (100% dichloromethane to 70%acetonitrile containing 3% methanol/30% dichloromethane) to give thedesired product (8.7 g, 62% for 2 steps) as a yellow foam as a 95:5mixture of enantiomers (RT=4.29 min and 6.00 min; Phenomenex LuxCellulose C-1 column, 4.6×150 mm, 5 micron particle size, eluting with15% ethanol in hexanes at 1 ml/min). This material was separated bychiral HPLC (Phenomenex Lux Cellulose C-1 column, 21.2×250 mm, 5 micronparticle size, eluting with 15% ethanol in hexanes at 10 ml/min) to give7.0 g of the desired peak 1 material (retention time of 8.20 min). ¹HNMR (300 MHz, CDCl₃) δ 8.24 (s, 1H), 7.51 (s, 1H), 6.32 (q, J=7.1 Hz,1H), 5.48 (br s, 2H), 4.40-4.18 (m, 5H), 4.05-3.93 (m, 1H), 3.81-3.65(m, 1H), 2.64 (s, 3H), 1.81 (d, J=7.1 Hz, 3H), 1.48 (t, J=7.0 Hz, 3H),1.43 (s, 9H). LCMS for C₂₅H₃₁ClN₇O₃(M+H)⁺: m/z=512.2; Found: 512.3.

Step 6.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile

A solution of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidine-1-carboxylate(peak 1 enantiomer from step 5) (2.2 g, 4.2 mmol) in methylene chloride(11 mL) was treated with trifluoroacetic acid (11 mL) dropwise andstirred at room temperature for 30 min. The reaction mixture wasconcentrated to an oil that was reconcentrated from ethanol (2×) to givea residue. This material was dissolved in a minimum amount of methanol,added dropwise to ice cooled saturated sodium bicarbonate solution (100ml), and extracted several times with 2:1 dichloromethane/isopropanol togive the desired product (1.8 g, quantitative) that was used withoutfurther purification. A small amount of the desired product was purifiedby preparative LCMS (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of60 mL/min) to give the desired product. The product was isolated as asingle enantiomer. ¹H NMR (400 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.47 (s,1H), 6.23 (q, J=7.0 Hz, 1H), 4.37-4.26 (m, 1H), 3.91-3.61 (m, 6H), 2.54(s, 3H), 1.71 (d, J=7.1 Hz, 3H), 1.32 (t, J=7.0 Hz, 3H). LCMS forC₂₀H₂₃ClN₇O (M+H)⁺: m/z=412.2; Found: 412.1.

Example 213.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(1-methylazetidin-3-yl)benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile(chiral intermediate in Example 212, Step 6) (0.30 g, 0.73 mmol) inmethanol (7.3 mL) was treated with formaldehyde (37% in water) (0.54 mL,7.3 mmol) and this was stirred at room temperature for 5 min. Thereaction mixture was treated with sodium cyanoborohydride (0.092 g, 1.5mmol) and stirred at room temperature for 2 h. The reaction mixture wasdiluted with methanol and purified by preparative LCMS (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 60 mL/min) to give the desiredproduct (0.16 g, 50%). The product was isolated as a single enantiomer.¹H NMR (400 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.48 (s, 1H), 6.27-6.18 (m,1H), 4.10-3.98 (m, 1H), 3.96-3.86 (m, 2H), 3.83-3.74 (m, 1H), 3.72-3.64(m, 1H), 3.10-2.98 (m, 2H), 2.54 (s, 3H), 2.20 (s, 3H), 1.71 (d, J=6.9Hz, 3H), 1.32 (t, J=6.7 Hz, 3H). LCMS for C₂₁H₂₅ClN₇O (M+H)⁺: m/z=426.2;Found: 426.2.

Example 219.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxyethyl)azetidin-3-yl]benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile(300 mg, 0.74 mmol, chiral intermediate from Example 212) intetrahydrofuran (14 mL) was treated with triethylamine (260 μL, 1.8mmol) followed by 2-bromoethanol (63 μL, 0.89 mmol) dropwise and stirredat 60° C. for 6 h. The reaction mixture was treated with additional2-bromoethanol (26 μL, 0.37 mmol) and stirred at 60° C. for another 6 h.The reaction mixture was poured into saturated sodium bicarbonatesolution and extracted with ethyl acetate. The organic layer wasconcentrated and purified by preparative LCMS (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 60 mL/min) to give the desired product (0.15g, 44%). The product was isolated as a single enantiomer. ¹H NMR (400MHz, DMSO-d₆) δ 8.19 (s, 1H), 7.56 (s, 1H), 6.36-6.25 (m, 1H), 4.48 (brs, 1H), 4.19-4.07 (m, 1H), 4.04-3.94 (m, 2H), 3.91-3.82 (m, 1H),3.81-3.72 (m, 1H), 3.20-3.08 (m, 2H), 2.62 (s, 2H), 2.57 (s, 3H), 1.79(d, J=6.8 Hz, 3H), 1.40 (t, J=6.6 Hz, 3H). LCMS for C₂₂H₂₇ClN₇O₂(M+H)⁺:m/z=456.2; Found: 456.1.

Example 220.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile(50 mg, 0.12 mmol, chiral intermediate from example 212) in ethanol (1.7mL) was treated with (S)-(−)-methyloxirane (21 μL, 0.30 mmol) and heatedin the microwave at 125° C. for 15 min. The reaction mixture was dilutedwith methanol and purified by preparative LCMS (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 60 mL/min) to give the desired product (27mg, 47%). The product was isolated as a single diastereomer. ¹H NMR (300MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.48 (s, 1H), 6.23 (q, J=6.9 Hz, 1H), 4.35(d, J=4.5 Hz, 1H), 4.13-3.99 (m, 1H), 3.97-3.88 (m, 2H), 3.85-3.63 (m,2H), 3.61-3.51 (m, 1H), 3.15-2.99 (m, 2H), 2.55 (s, 3H), 2.28 (d, J=5.9Hz, 2H), 1.71 (d, J=7.0 Hz, 3H), 1.32 (t, J=6.9 Hz, 3H), 1.00 (d, J=6.2Hz, 3H). LCMS for C₂₃H₂₉ClN₇O₂(M+H)⁺: m/z=470.2; Found: 470.2.

Example 236. tert-Butyl2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoate

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile(0.38 g, 0.92 mmol, chiral intermediate from Example 212) inN,N-dimethylformamide (4. 6 mL) was treated with potassium carbonate(0.51 g, 3.7 mmol) followed by tert-butyl 2-bromo-2-methylpropanoate(0.86 mL, 4.6 mmol) and heated at 60° C. for 3 h. The reaction mixturewas poured into water and extracted with ethyl acetate. The organiclayer was separated, dried with magnesium sulfate, filtered, andconcentrated to a crude oil. The crude material was purified by flashcolumn chromatography using methanol in dichloromethane (0%-10%) to givethe desired product (0.43 g, 83%). The product was isolated as a singleenantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.10 (s, 1H), 7.44 (s, 1H), 6.22(q, J=6.8 Hz, 1H), 4.12-3.97 (m, 1H), 3.88-3.70 (m, 4H), 3.62-3.48 (m,2H), 2.54 (s, 3H), 1.70 (d, J=7.0 Hz, 3H), 1.33 (t, J=6.9 Hz, 3H), 1.17(s, 9H), 1.05 (s, 6H). LCMS for C₂₈H₃₇ClN₇O₃(M+H)⁺: m/z=554.3; Found:554.3.

Example 237.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxy-1,1-dimethylethyl)azetidin-3-yl]benzonitrile

Step 1.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoicacid bis(trifluoroacetate)

tert-Butyl2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoate(0.36 g, 0.65 mmol, chiral intermediate from Example 236) was dissolvedin a premixed solution of trifluoroacetic acid (3.2 mL)/water (0.065 mL)and stirred at room temperature for 3 h and at 50° C. for 30 min. Thereaction mixture was concentrated and reconcentrated from acetonitrile(2×) to give the desired product as a gum. This gum was treated with asmall amount of methyl-tert-butylether that was swirled until a solidformed. The methyl-tert-butylether was decanted and the residue wasconcentrated to give the desired product (0.51 g, 109%) that was usedwithout further purification. LCMS for C₂₄H₂₉ClN₇O₃(M+H)⁺: m/z=498.2;Found: 498.3.

Step 2.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxy-1,1-dimethylethyl)azetidin-3-yl]benzonitrile

A solution of2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoicacid bis(trifluoroacetate) (0.10 g, 0.16 mmol) in tetrahydrofuran (0.9mL) was cooled to −25° C., treated with 4-methylmorpholine (0.072 mL,0.65 mmol) and isobutyl chloroformate (0.085 mL, 0.65 mmol), and stirredat −15° C. for 15 min. The reaction mixture was filtered though adisposable filter cartridge into a separate round bottom flask. Thissolution was then cooled to −20° C. and a solution of sodiumtetrahydroborate (0.031 g, 0.82 mmol) in a minimum amount of water wasadded dropwise. The reaction mixture was stirred at −15° C. for 30 min,poured into water, and extracted with ethyl acetate. The organic layerwas separated, concentrated, diluted with methanol, and purified bypreparative LCMS (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) to give the desired product (3.5 mg, 4%). The product wasisolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s,1H), 7.50 (s, 1H), 7.35 (br s, 2H), 6.23 (q, J=6.7 Hz, 1H), 4.44-4.35(m, 1H), 4.04-3.88 (m, 1H), 3.86-3.73 (m, 1H), 3.72-3.57 (m, 3H), 3.12(d, J=4.7 Hz, 2H), 2.54 (s, 3H), 1.71 (d, J=6.9 Hz, 3H), 1.31 (t, J=6.9Hz, 3H), 0.80 (s, 6H). LCMS for C₂₄H₃₁ClN₇O₂(M+H)⁺: m/z=484.2; Found:484.2.

Example 239.2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanamide

A solution of2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoicacid bis(trifluoroacetate) (0.05 g, 0.069 mmol, chiral intermediate fromExample 237, Step 1) and 2.0 M ammonia in ethanol (0.17 mL, 0.34 mmol)in N,N-dimethylformamide (1 mL) was treated with triethylamine (0.048mL, 0.35 mmol) and benzotriazol-1-yloxytris(dimethylamino)phosphoniumhexafluorophosphate (0.046 g, 0.10 mmol) and stirred at room temperaturefor 1 h. The reaction mixture was quenched with a few drops of water,diluted with methanol, and purified by preparative LCMS (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 60 mL/min) to give the desiredproduct (25 mg, 73%). The product was isolated as a single enantiomer.¹H NMR (400 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.51 (s, 1H), 7.23 (s, 1H),6.98 (s, 1H), 6.23 (q, J=7.0 Hz, 1H), 4.09-3.96 (m, 1H), 3.84-3.61 (m,4H), 3.39-3.34 (m, 1H), 3.32-3.28 (m, 1H), 2.54 (s, 3H), 1.71 (d, J=7.0Hz, 3H), 1.31 (t, J=6.9 Hz, 3H), 1.02 (s, 6H). LCMS forC₂₄H₃₀ClN₈O₂(M+H)⁺: m/z=497.2; Found: 497.3.

Example 247.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxy-2-methylpropanoyl)azetidin-3-yl]benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile(0.04 g, 0.097 mmol, chiral intermediate from Example 212) and propanoicacid, 2-hydroxy-2-methyl- (0.012 g, 0.12 mmol) in N,N-dimethylformamide(0.54 mL) was treated with triethylamine (0.034 mL, 0.24 mmol) followedby O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (0.048 g, 0.13 mmol) and stirred at room temperaturefor 30 min. The reaction mixture was diluted with methanol andacetonitrile and purified by preparative LCMS (XBridge C18 column,eluting with a gradient of methanol/water containing 0.1% ammoniumhydroxide, at flow rate of 60 mL/min) to give the desired product (7 mg,14%). The product was isolated as a single enantiomer. ¹H NMR (300 MHz,DMSO-d₆) δ 8.11 (s, 1H), 7.54 (d, J=4.5 Hz, 1H), 6.25 (q, J=7.2 Hz, 1H),5.08 (s, 1H), 4.88-4.77 (m, 1H), 4.73-4.60 (m, 1H), 4.50-4.35 (m, 1H),4.29-4.09 (m, 2H), 3.85-3.73 (m, 2H), 2.55 (s, 3H), 1.73 (d, J=7.0 Hz,3H), 1.37 (t, J=6.3 Hz, 3H), 1.26 (s, 3H), 1.22 (s, 3H). LCMS forC₂₄H₂₉ClN₇O₃(M+H)⁺: m/z=498.2; Found: 498.2.

Example 261.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile

Step 1. 4-Acetyl-6-chloro-2-iodo-3-methoxybenzonitrile

A solution of 1-(5-chloro-4-fluoro-3-iodo-2-methoxyphenyl)ethanone(intermediate from Example 13, Step 3) (18 g, 54 mmol) inN,N-dimethylformamide (200 mL) was treated with potassium cyanide (5.2g, 81 mmol) and stirred at 40° C. for 6 h. The reaction mixture wasdiluted with ethyl acetate and poured into saturated sodium bicarbonatesolution/water (1:1). The organic layer was separated, washed withsaturated sodium bicarbonate solution, dried with magnesium sulfate,filtered, and concentrated to give a crude brown oil. The crude materialwas purified by flash column chromatography using ethyl acetate inhexanes (0%-30%) to give the desired product (11 g, 61%) as a yellowsolid. ¹H NMR (300 MHz, CDCl₃) δ 7.60 (s, 1H), 3.81 (s, 3H), 2.62 (s,3H). LCMS for C₁₀H₈ClINO₂ (M+H)⁺: m/z=335.9; Found: 335.9.

Step 2. tert-Butyl3-(3-acetyl-5-chloro-6-cyano-2-methoxyphenyl)azetidine-1-carboxylate

Zinc (5.0 g, 77 mmol) and oven dried Celite (520 mg) was added to aflask and the flask was heated with a heat gun while under high-vac for5 min and then back-filled with nitrogen. N, N-dimethylacetamide (53 mL)was added, followed by 1,2-dibromoethane (400 μL, 4.6 mmol) and themixture was heated at 70° C. for 15 min and then cooled to roomtemperature. The reaction mixture was treated with chlorotrimethylsilane(580 μL, 4.6 mmol) dropwise and stirred at room temperature for 1 h. Thereaction mixture was treated with a solution of tert-butyl3-iodoazetidine-1-carboxylate (16 g, 58 mmol) in N,N-dimethylacetamide(26 mL) dropwise (internal temperature was kept below 40° C. with awater bath) and heated at 40° C. for 2 h. The zinc-iodo reagent(transferred via canula) was filtered through a plastic filter (that wasappropriately sealed to avoid atmospheric exposure) directly into aclean, dry flask that was flushed with nitrogen. The reaction mixturewas treated with tris(dibenzylideneacetone)dipalladium(0) (670 mg, 0.73mmol) and tri-(2-furyl)phosphine (340 mg, 1.5 mmol) and degassed withnitrogen for a few minutes. The reaction mixture was treated with asolution of 4-acetyl-6-chloro-2-iodo-3-methoxybenzonitrile (13 g, 39mmol) in N,N-dimethylacetamide (120 mL) (degassed with nitrogen) quicklyand heated at 70° C. for 2 h. The reaction mixture was poured intosaturated ammonium chloride solution and extracted with ethyl acetate(3×300 mL). The combined organic extracts were washed with water (4×500mL) and brine (1×500 mL), dried with magnesium sulfate, filtered, andconcentrated to a crude dark oil. The crude material was purified byflash column chromatography using ethyl acetate in hexanes (5%-40%) togive the desired product (12 g, 85%). ¹H NMR (400 MHz, DMSO-d₆) δ 7.79(s, 1H), 4.39-4.29 (m, 1H), 4.28-4.11 (m, 4H), 3.68 (s, 3H), 2.58 (s,3H), 1.38 (s, 9H).

Step 3. tert-Butyl3-[3-chloro-2-cyano-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate

A solution of(3aS)-1-methyl-3,3-diphenyltetrahydro-3H-pyrrolo[1,2-c][1,3,2]oxazaborole(4.3 g, 16 mmol) in tetrahydrofuran (46 mL) was treated with 1.0 Mborane-THF complex in tetrahyrofuran (19 mL, 19 mmol) and stirred at 20°C. for 15 min. The reaction mixture was cooled to −30° C. and treatedwith a solution of tert-butyl3-(3-acetyl-5-chloro-6-cyano-2-methoxyphenyl)azetidine-1-carboxylate(5.7 g, 16 mmol) in tetrahydrofuran (49 mL) slowly. The flask containingthe starting material ketone was rinsed with additional tetrahydrofuran(9 mL) and added to the reaction mixture. The temperature of thereaction was −20° C. after the addition was complete. The reactionmixture was warmed to −5° C. over a period of 30 min. The reactionmixture was quenched with water at 0° C., poured into saturated sodiumbicarbonate solution, and extracted with ethyl acetate. The aqueouslayer was separated and extracted with ethyl acetate. The combinedorganic layers were washed with water and brine, dried with magnesiumsulfate, filtered, and concentrated to a crude dark oil. The crudematerial was purified by flash column chromatography using ethyl acetatein hexanes (0%-100%) to give the desired product (5.5 g, 97%) as a beigefoam as a 97:3 mixture of enantiomers (Retention times=12.19 min and13.18 min; Phenomenex Lux Cellulose C-2 column, 4.6×150 mm, 5 micronparticle size, eluting with 8% ethanol in hexanes at 1 ml/min). ¹H NMR(400 MHz, DMSO-d₆) δ 7.62 (s, 1H), 5.48 (d, J=4.6 Hz, 1H), 5.00-4.90 (m,1H), 4.43-4.31 (m, 1H), 4.30-4.10 (m, 4H), 3.66 (s, 3H), 1.38 (s, 9H),1.29 (d, J=6.4 Hz, 3H). LCMS for C₁₄H₁₆ClN₂O₄ ([M-(t-Bu)+H]+H)⁺:m/z=311.1; Found: 311.1.

Step 4. tert-Butyl3-{3-[1-(4-amino-3-methyl-H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxylate

A solution of tert-butyl3-[3-chloro-2-cyano-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate(8.6 g, 23 mmol) (97:3 mixture of enantiomers from step 3) in methylenechloride (220 mL) at 0° C. was treated with triethylamine (8.2 mL, 59mmol) followed by methanesulphonic anhydride (6.1 g, 35 mmol) andstirred at 0° C. for 15 min. The reaction mixture was diluted withdichloromethane and washed with water and brine, dried with magnesiumsulfate, filtered, and concentrated to give the crude mesylate that wasused without further purification. A solution of the crude mesylateintermediate in N,N-dimethylformamide (82 mL) was cooled to 0° C.,treated with sodium hydride (1.2 g, 30 mmol) (60% in mineral oil), andstirred at 0° C. for 30 min. The reaction mixture was treated with asolution of tert-butyl3-(3-chloro-2-cyano-6-methoxy-5-{1-[(methylsulfonyl)oxy]ethyl}phenyl)azetidine-1-carboxylate(11 g, 24 mmol) in N,N-dimethylformamide (170 mL) dropwise over a periodof 10 min and stirred at 0° C. for 30 min and heated at 50° C. for 1 h.The reaction mixture was diluted with water and saturated sodiumbicarbonate solution and extracted with ethyl acetate (3×200 mL). Thecombined organic extracts were washed with water (4×150 mL) and brine,dried with magnesium sulfate, filtered, and concentrated to a crude oil.The crude material was purified by flash column chromatography (2%methanol/98% dichloromethane to 7% methanol/93% dichloromethane [thedichloromethane contained 0.5% triethylamine]) to give the desiredproduct (9.1 g, 77% for 2 steps) as a 9:1 mixture of enantiomers. Thismaterial was separated by chiral HPLC (retention times=5.81 min and 8.94min; Chiracel AD-H column, 20×250 mm, 5 micron particle size, elutingwith 10% ethanol in hexanes at 18 ml/min, 10 mg/inj) to give 6.9 g ofthe desired peak 1 material. ¹H NMR (400 MHz, DMSO-d₆) δ 8.11 (s, 1H),7.52 (s, 1H), 6.25 (q, J=7.0 Hz, 1H), 4.45-4.33 (m, 1H), 4.27-4.13 (m,4H), 3.70 (s, 3H), 2.55 (s, 3H), 1.73 (d, J=7.1 Hz, 3H), 1.37 (s, 9H).LCMS for C₂₀H₂₁ClN₇O₃ ([M-(t-Bu)+H]+H)⁺: m/z=442.1; Found: 442.1.

Step 5.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile

A solution of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxylate(1.7 g, 3.3 mmol) in methylene chloride (30 mL) was treated withtrifluoroacetic acid (20 mL) and stirred at room temperature for 20 min.The reaction mixture was concentrated to give a residue that was dilutedwith methanol (50 mL) and saturated sodium bicarbonate solution (50 mL).This aqueous solution was diluted with brine (50 mL) and extracted witha 5:1 mixture of dichloromethane/isopropanol (5×100 mL). The combinedorganic extracts were dried over sodium sulfate and concentrated to givethe desired product (1.4 g, 97%). The product was isolated as a singleenantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.46 (s, 1H), 7.34(br s, 2H), 6.24 (q, J=6.9 Hz, 1H), 4.40-4.26 (m, 1H), 3.90-3.68 (m,4H), 3.63 (s, 3H), 2.55 (s, 3H), 1.72 (d, J=7.1 Hz, 3H). LCMS forC₁₉H₂₁ClN₇O (M+H)⁺: m/z=398.1; Found: 398.1.

Example 262.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxy-2-(1-methylazetidin-3-yl)benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (50 mg, 0.13 mmol) in methanol (3mL) was treated with sodium cyanoborohydride (20 mg, 0.31 mmol) followedby formaldehyde (37% in water) (37 μL, 0.50 mmol) and stirred at roomtemperature for 20 min. The reaction mixture was quenched with aceticacid (170 μL, 2.9 mmol), diluted with methanol, and purified bypreparative LCMS (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of60 mL/min) to give the desired product (30 mg, 58%). The product wasisolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s,1H), 7.46 (s, 1H), 7.37 (br s, 2H), 6.23 (q, J=7.0 Hz, 1H), 4.10-3.96(m, 1H), 3.95-3.85 (m, 2H), 3.63 (s, 3H), 3.05-2.94 (m, 2H), 2.55 (s,3H), 2.18 (s, 3H), 1.72 (d, J=7.1 Hz, 3H). LCMS for C₂₀H₂₃ClN₇O (M+H)⁺:m/z=412.2; Found: 412.1.

Example 268.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-[1-(2-hydroxyethyl)azetidin-3-yl]-3-methoxybenzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (400 mg, 1.0 mmol) intetrahydrofuran (14 mL) was treated with triethylamine (350 μL, 2.5mmol) and 2-bromoethanol (85 μL, 1.2 mmol) and stirred at 60° C.overnight. The reaction mixture was concentrated, diluted with methanol,and purified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired product (0.14 g, 31%). Theproduct was isolated as a single enantiomer. ¹H NMR (400 MHz, DMSO-d₆) δ8.11 (s, 1H), 7.46 (s, 1H), 6.24 (q, J=6.9 Hz, 1H), 4.41 (t, J=5.4 Hz,1H), 4.12-4.03 (m, 1H), 3.97-3.88 (m, 2H), 3.64 (s, 3H), 3.38-3.34 (m,2H), 3.09-3.01 (m, 2H), 2.55 (s, 3H), 2.41 (t, J=5.9 Hz, 2H), 1.72 (d,J=7.0 Hz, 3H). LCMS for C₂₁H₂₅ClN₇O₂(M+H)⁺: m/z=442.2; Found: 442.2.

The compounds of Example 268 and 269 were synthesized from the samechiral intermediate in Example 261. According to the crystal structuredetermination in Example 269, the stereochemistry at the carbon at the1-position of the ethan-1,1-diyl group is S. Because the compound ofExample 268 was synthesized from the same chiral intermediate as Example269, one of ordinary skill in the art would expect that the carbon atthe 1-position of the ethan-1,1-diyl group of Example 268 is also in theS-configuration. Accordingly, it is believed that the compound ofExample 268 is(S)-4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-(2-hydroxyethyl)azetidin-3-yl)-3-methoxybenzonitrile.

Example 269.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}-3-methoxybenzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (2.5 g, 6.3 mmol) in ethanol (130mL) was treated with (S)-(−)-methyloxirane (1.1 mL, 16 mmol) and heatedin the microwave at 120° C. for 25 min. The reaction mixture wasconcentrated to give a residue that was purified by flash columnchromatography using methanol in dichloromethane (0%-10%; methanolcontained 0.5% triethylamine) and by preparative LCMS (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 60 mL/min) to give the desiredproduct (0.76 g, 26%). The product was isolated as a singlediastereomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.46 (s, 1H),7.34 (br s, 2H), 6.23 (q, J=7.0 Hz, 1H), 4.35 (br s, 1H), 4.14-3.99 (m,1H), 3.98-3.87 (m, 2H), 3.64 (s, 3H), 3.60-3.52 (m, 1H), 3.13-2.99 (m,2H), 2.55 (s, 3H), 2.28 (d, J=5.9 Hz, 2H), 1.75-1.69 (m, 3H), 1.00 (d,J=6.2 Hz, 3H). LCMS for C₂₂H₂₇ClN₇O₂(M+H)⁺: m/z=456.2; Found: 456.2.

Crystal Structure Determination for the Compound of Example 269C22,H26,N7,O2,CL1+H₂O

CRYSTAL DATA: C22 H28 Cl F0 N7 O3, from ACN/water, colorless, needle,˜0.500×0.070×0.050 mm, monoclinic, C2, a=25.941(7) Å, b=4.9767(13) Å,c=17.787(5) Å, beta=101.967(4)°, Vol=2246.3(10) Å³, Z=4, T=−100° C.,Formula weight=473.96, Density=1.401 g/cm³, (Mo)=0.21 mm⁻¹DATA COLLECTION: Bruker SMART APEX-II CCD system, MoKalpha radiation,standard focus tube, anode power=50 kV×42 mA, crystal to platedistance=5.0 cm, 512×512 pixels/frame, beam center=(256.13, 253.14),total frames=704, oscillation/frame=0.50°, exposure/frame=120.1sec/frame, SAINT integration, hkl min/max=(−27, 34, −6, 6, −23, 11),data input to shelx=7578, unique data=5186, two-theta range=3.20 to56.740, completeness to two-theta 56.74=99.70%, R(int-xl)=0.0331, SADABScorrection applied.SOLUTION AND REFINEMENT: Structure solved using XS(Shelxtl), refinedusing shelxtl software package, refinement by full-matrix least squareson F², scattering factors from Int. Tab. Vol C Tables 4.2.6.8 and6.1.1.4, number of data=5186, number of restraints=2, number ofparameters=313, data/parameter ratio=16.57, goodness-of-fit on F²=1.02,R indices[I>4sigma(I)] R1=0.0524, wR2=0.1033, R indices(all data)R1=0.0826, wR2=0.1162, max difference peak and hole=0.294 and −0.221e/Å³, refined flack parameter=0.05(8), All of the hydrogen atoms exceptthe NH2 and water hydrogens have been idealized using a riding model.RESULTS: The asymmetric unit contains one molecule and one watermolecule as shown in FIG. 1 with thermal ellipsoids drawn to the 50%probability level. The predicted structure is confirmed. The absoluteconfiguration is determined based upon the known S configuration at C21.The configuration at C7 is determined to be S. The flack parameter alsoconfirms the correct configuration. Based on the crystal structure, thecompound of Example 269 is believed to be4-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-((S)-2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile.The crystal structure is shown in FIG. 1.

Examples 272 and 273. Diastereoisomers of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-[1-(2-hydroxy-1-methylethyl)azetidin-3-yl]-3-methoxybenzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(40 mg, 0.10 mmol) in methanol (2 mL) was treated with sodiumcyanoborohydride (16 mg, 0.25 mmol) followed by acetol (28 μL, 0.40mmol) and stirred at room temperature for 1 h. The reaction mixture wasquenched with acetic acid (100 μL, 1.8 mmol), diluted with methanol, andpurified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired products as a mixture ofdiastereoisomers. This mixture of diastereoisomers was separated bychiral HPLC (RT=3.70 min and 6.58 min; Phenomenex Lux Cellulose C-4column, 21.2×250 mm, 5 micron particle size, eluting with 20% ethanol inhexanes at 18 ml/min, 5 mg/inj) to give the desired peak 1 isomer(compound 272) (19 mg, 41%) and peak 2 isomer (compound 273) (23 mg,50%) Peak 1: ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.47 (s, 1H),7.34 (br s, 2H), 6.24 (q, J=6.9 Hz, 1H), 4.43 (t, J=5.2 Hz, 1H),4.07-3.82 (m, 3H), 3.64 (s, 3H), 3.31-3.24 (m, 1H), 3.17-3.06 (m, 2H),3.06-2.97 (m, 1H), 2.55 (s, 3H), 2.21-2.11 (m, 1H), 1.72 (d, J=7.1 Hz,3H), 0.81 (d, J=6.3 Hz, 3H). LCMS for C₂₂H₂₇ClN₇O₂(M+H)⁺: m/z=456.2;Found: 456.2. Peak 2: ¹H NMR (300 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.47 (s,1H), 7.35 (br s, 2H), 6.24 (q, J=7.0 Hz, 1H), 4.43 (t, J=5.5 Hz, 1H),4.06-3.91 (m, 2H), 3.89-3.79 (m, 1H), 3.64 (s, 3H), 3.30-3.24 (m, 1H),3.15-3.00 (m, 3H), 2.55 (s, 3H), 2.21-2.10 (m, 1H), 1.72 (d, J=7.1 Hz,3H), 0.82 (d, J=6.2 Hz, 3H). LCMS for C₂₂H₂₇ClN₇O₂(M+H)⁺: m/z=456.2;Found: 456.2.

Example 281.2-(1-Acetylazetidin-3-yl)-4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxybenzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (60 mg, 0.15 mmol) intetrahydrofuran (2 mL) at 0° C. was treated with triethylamine (53 μL,0.38 mmol) followed by acetyl chloride (13 μL, 0.18 mmol) and stirred at20° C. overnight. The reaction mixture was diluted with methanol andpurified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired product (39 mg, 59%). Theproduct was isolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ8.11 (s, 1H), 7.52 (d, J=2.5 Hz, 1H), 7.36 (br s, 2H), 6.26 (q, J=7.0Hz, 1H), 4.57-4.36 (m, 3H), 4.30-4.21 (m, 1H), 4.18-4.08 (m, 1H), 3.71(d, J=3.1 Hz, 3H), 2.55 (s, 3H), 1.78-1.71 (m, 6H). LCMS forC₂₁H₂₃ClN₇O₂(M+H)⁺: m/z=440.2; Found: 440.1.

Example 285.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxy-2-[1-(methylsulfonyl)azetidin-3-yl]benzonitrile

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (40 mg, 0.10 mmol) indichloromethane (1 mL) was treated with triethylamine (35 μL, 0.25mmol), cooled to 0° C., treated with methanesulfonyl chloride (9.3 μL,0.12 mmol) and stirred at 0° C. for 1 h. The reaction mixture wasdiluted with methanol and purified by preparative LCMS (XBridge C18column, eluting with a gradient of acetonitrile/water containing 0.1%ammonium hydroxide, at flow rate of 60 mL/min) to give the desiredproduct (20 mg, 42%). The product was isolated as a single enantiomer.¹H NMR (300 MHz, DMSO-d₆) δ 8.12 (s, 1H), 7.55 (s, 1H), 7.35 (br s, 2H),6.25 (q, J=7.0 Hz, 1H), 4.54-4.40 (m, 1H), 4.27-4.12 (m, 4H), 3.68 (s,3H), 3.01 (s, 3H), 2.55 (s, 3H), 1.74 (d, J=7.1 Hz, 3H). LCMS forC₂₀H₂₃ClN₇O₃S (M+H)⁺: m/z=476.1; Found: 476.1.

Example 289. Methyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxylate

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (20 mg, 0.05 mmol) indichloromethane (1 mL) was treated with triethylamine (20 μL, 0.14 mmol)followed by methyl chloroformate (4.7 μL, 0.06 mmol) and stirred at roomtemperature for 1 h. The reaction mixture was diluted with methanol andpurified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired product (12 mg, 52%). Theproduct was isolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ8.11 (s, 1H), 7.51 (s, 1H), 7.34 (br s, 2H), 6.25 (q, J=7.0 Hz, 1H),4.53-4.38 (m, 1H), 4.36-4.17 (m, 4H), 3.71 (s, 3H), 3.55 (s, 3H), 2.55(s, 3H), 1.73 (d, J=7.1 Hz, 3H). LCMS for C₂₁H₂₃ClN₇O₃(M+H)⁺: m/z=456.2;Found: 456.1.

Example 292.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}-N-(tert-butyl)azetidine-1-carboxamide

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (20 mg, 0.05 mmol) inN,N-dimethylformamide (1 mL) was treated with triethylamine (20 μL, 0.14mmol) followed by 2-isocyanato-2-methyl-propane (7.2 μL, 0.063 mmol) andstirred at room temperature overnight. The reaction mixture was dilutedwith methanol and purified by preparative LCMS (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 60 mL/min) to give the desired product (16mg, 64%). The product was isolated as a single enantiomer. LCMS forC₂₄H₃₀ClN₈O₂(M+H)⁺: m/z=497.2; Found: 497.2.

Example 293.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxamide

A solution of3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}-N-(tert-butyl)azetidine-1-carboxamide(chiral intermediate from Example 292) (16 mg, 0.032 mmol) intrifluoroacetic acid (2 mL) was heated in the microwave at 120° C. for10 min. The reaction mixture was diluted with methanol and purified bypreparative LCMS (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of60 mL/min) to give the desired product (7 mg, 50%). The product wasisolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ 8.12 (s,1H), 7.62 (s, 1H), 7.35 (br s, 2H), 6.28 (q, J=6.9 Hz, 1H), 5.70 (br s,1H), 4.62-4.49 (m, 1H), 4.34-4.20 (m, 1H), 3.83 (s, 3H), 3.78-3.49 (m,2H), 2.55 (s, 3H), 1.73 (d, J=7.0 Hz, 3H). LCMS for C₂₀H₂₂ClN₈O₂(M+H)⁺:m/z=441.2; Found: 441.1.

Example 296.3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}-N,N-dimethylazetidine-1-carboxamide

A solution of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile(chiral intermediate from Example 261) (40 mg, 0.10 mmol) inN,N-dimethylformamide (2 mL) was treated with triethylamine (40 μL, 0.29mmol) followed by p-nitrophenyl chloroformate (23 μL, 0.13 mmol) andstirred at room temperature for 1 h. The reaction mixture was dilutedwith methanol and purified by preparative LCMS (XBridge C18 column,eluting with a gradient of acetonitrile/water containing 0.1% ammoniumhydroxide, at flow rate of 60 mL/min) to give the desired product thatwas used immediately. A solution of the p-nitrophenyl carbamateintermediate in tetrahydrofuran (1 mL) was treated with triethylamine(15 μL, 0.11 mmol) followed by a solution of 1.0 M dimethylamine intetrahydrofuran (150 μL, 0.15 mmol) and heated in a sealed tube at 60°C. for 2 h. The reaction mixture was concentrated, diluted with methanoland purified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired product (13 mg, 28%). Theproduct was isolated as a single enantiomer. ¹H NMR (300 MHz, DMSO-d₆) δ8.11 (s, 1H), 7.49 (s, 1H), 7.36 (br s, 2H), 6.25 (q, J=7.0 Hz, 1H),4.44-4.23 (m, 3H), 4.22-4.10 (m, 2H), 3.69 (s, 3H), 2.76 (s, 6H), 2.55(s, 3H), 1.73 (d, J=7.1 Hz, 3H). LCMS for C₂₂H₂₆ClN₈O₂(M+H)⁺: m/z=469.2;Found: 469.1.

Example 298.1-{1-[4,5-Dichloro-3-(1-ethylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

Step 1. 1-(4,5-Dichloro-2-hydroxyphenyl)ethanone

A solution of 3,4-dichlorophenol [AK Scientific] (30 g, 18 mmol) inacetyl chloride (19 mL, 270 mmol) was stirred at 60° C. for 2 h. Thereaction mixture was cooled to 20° C., treated with aluminum trichloride(37 g, 280 mmol) portionwise, and heated at 180° C. for 30 min. Thereaction mixture was cooled to 20° C. and the solution hardened into asolid block that was not easy to break apart. This material was cooledto 0° C. and quenched slowly with 1 M HCl in portions. The solid blockof material slowly broke apart with enough HCl and this heterogenousmixture was stirred at 20° C. overnight to ensure uniformity. The solidwas filtered, washed with copious amounts of water, and dried undervacuum to give the desired product (38 g, quantitative) as a tan solid.

Step 2. 1-(4,5-Dichloro-2-hydroxy-3-iodophenyl)ethanone

A solution of 1-(4,5-dichloro-2-hydroxyphenyl)ethanone (12 g, 59 mmol)in acetic acid (70 mL) was treated with N-iodosuccinimide (16 g, 71mmol) and stirred at 90° C. for 18 h. The reaction mixture was treatedwith additional N-iodosuccinimide (8 g, 36 mmol) and stirred at 90° C.for 4 h. The reaction mixture was concentrated, diluted with ethylacetate, and quenched with saturated sodium bicarbonate until thebubbling stopped. The organic layer was separated and the aqueous wasre-extracted with ethyl acetate. The combined organic layers were driedand concentrated to give a brown solid. This material was recrystallizedfrom methanol to give desired product (9.0 g, 46%) as a tan solid. ¹HNMR (300 MHz, CDCl₃) δ 13.36 (s, 1H), 7.85 (s, 1H), 2.65 (s, 3H). LCMSfor C₈H₆Cl₂IO₂ (M+H)⁺: m/z=330.9, 332.9; Found: 330.8, 332.9.

Step 3. 1-(4,5-Dichloro-3-iodo-2-methoxyphenyl)ethanone

A solution of 1-(4,5-dichloro-2-hydroxy-3-iodophenyl)ethanone (16 g, 47mmol) and potassium carbonate (17 g, 120 mmol) in N,N-dimethylformamide(40 mL) was treated with methyl iodide (6.4 mL, 100 mmol) and stirred at60° C. for 1 h. The reaction mixture was diluted with water andextracted with ethyl acetate (2×). The combined organic layers weredried with magnesium sulfate, filtered, and concentrated to give a crudesolid. The crude material was purified by flash column chromatographyusing ethyl acetate in hexanes (5%-30%) to give the desired product (14g, 84%) as an orange solid. ¹H NMR (300 MHz, CDCl₃) δ 7.69 (s, 1H), 3.79(s, 3H), 2.60 (s, 3H). LCMS for C₉H₈Cl₂IO₂ (M+H)⁺: m/z=344.9, 346.9;Found: 344.8, 346.9.

Step 4. tert-Butyl3-(3-acetyl-5,6-dichloro-2-methoxyphenyl)azetidine-1-carboxylate

Zinc (4.5 g, 69 mmol) was suspended with 1,2-dibromoethane (420 μL, 4.9mmol) in N, N-dimethylformamide (54 mL). The mixture was heated at 70°C. for 10 min and then cooled to room temperature. Chlorotrimethylsilane(620 μL, 4.9 mmol) was added dropwise and stirring was continued for 1h. A solution of tert-butyl 3-iodoazetidine-1-carboxylate (17 g, 61mmol) in N,N-dimethylformamide (30 mL) was then added and the mixturewas heated at 40° C. for 1 h before a mixture of1-(4,5-dichloro-3-iodo-2-methoxyphenyl)ethanone (14 g, 41 mmol),tris(dibenzylideneacetone)dipalladium(0) (710 mg, 0.77 mmol) andtri-(2-furyl)phosphine (360 mg, 1.6 mmol) in N,N-dimethylformamide (120mL) was added quickly. The reaction mixture was stirred overnight atroom temperature. The reaction mixture was then partitioned betweenethyl acetate and saturated ammonium chloride solution. The organiclayer was washed with water, dried with magnesium sulfate, filtered, andconcentrated to a crude residue that was purified by flash columnchromatography using ethyl acetate in hexanes (0%-25%) to give thedesired product (12 g, 77%). LCMS for C₁₇H₂₁Cl₂NO₄Na (M+Na)⁺: m/z=396.1;Found: 396.0.

Step 5. tert-Butyl3-[2,3-dichloro-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate

A solution of tert-butyl3-(3-acetyl-5,6-dichloro-2-methoxyphenyl)azetidine-1-carboxylate (9.6 g,26 mmol) in methanol (240 mL) at 0° C. was treated with sodiumtetrahydroborate (1.9 g, 51 mmol) portionwise over 5 min and stirred at0° C. for 30 min. The reaction mixture was quenched with acetic acid(7.3 mL, 130 mmol) at 0° C. and treated with saturated sodiumbicarbonate solution (˜50 mL). The reaction mixture was concentrated toremove most of the methanol (to ˜60 mL), poured into saturated sodiumbicarbonate solution (150 ml), and extracted with ethyl acetate (2×200mL). The combined organic extracts were washed with water and brine,dried over sodium sulfate, filtered, and concentrated to give thedesired product (9.6 g, quantitative) that was used without furtherpurification. LCMS for C₁₃H₁₆Cl₂NO₄ ([M-(t-Bu)+H]+H)⁺: m/z=320.0; Found:320.0.

Step 6. tert-Butyl3-[2,3-dichloro-5-(1-chloroethyl)-6-methoxyphenyl]azetidine-1-carboxylate

N,N-Dimethylformamide (0.92 mL, 12 mmol) was added to solid cyanuricchloride (2.2 g, 12 mmol) at room temperature (DMF is absorbed by thesolid). The mixture was allowed to stand for 10 min, treated withmethylene chloride (60 mL), and stirred for a few minutes to break upthe solid. The reaction mixture was treated with a solution oftert-butyl3-[2,3-dichloro-5-(1-hydroxyethyl)-6-methoxyphenyl]azetidine-1-carboxylate(3.0 g, 8.0 mmol) in methylene chloride (30 mL) and stirred at 35-40° C.for 2 h. The reaction mixture was treated with additionalN,N-dimethylformamide (1 mL) and stirred at 35-40° C. for 4 h. Thereaction required another treatment of N,N-dimethylformamide (1 mL) withstirring at 35-40° C. overnight to proceed to completion. The reactionmixture was diluted with water and dichloromethane. The organic phasewas separated and washed with saturated sodium bicarbonate solution,water and brine, dried over magnesium sulfate, filtered, andconcentrated to a crude residue. The crude material was purified byflash column chromatography using ethyl acetate in hexanes (5%-40%) togive the desired product (2.8 g, 90%). LCMS for C₁₃H₁₅Cl₃NO₃([M-(t-Bu)+H]+H)⁺: m/z=338.0, 340.0; Found: 337.9, 339.9.

Step 7. tert-Butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl)ethyl]-5,6-dichloro-2-methoxyphenyl}azetidine-1-carboxylate

A solution of tert-butyl3-[2,3-dichloro-5-(1-chloroethyl)-6-methoxyphenyl]azetidine-1-carboxylate(1.0 g, 2.5 mmol) and 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.43g, 2.9 mmol) in N,N-dimethylformamide (23 mL) was treated with cesiumcarbonate (1.2 g, 3.8 mmol) and potassium iodide (42 mg, 0.25 mmol) andheated at 100° C. for 10 h. The reaction mixture was diluted with ethylacetate (75 mL) and water (75 mL). The aqueous layer was separated andreextracted with ethyl acetate (2×50 mL). The combined organic layerswere washed with water, saturated sodium bicarbonate solution, andbrine, dried over magnesium sulfate, filtered, and concentrated to acrude residue. The crude material was purified by flash columnchromatography using methanol in dichloromethane (0%-10%) to give thedesired product (0.97 g, 75%). LCMS for C₂₃H₂₉Cl₂N₆O₃ (M+H)⁺: m/z=507.2,509.2; Found: 507.0, 509.0.

Step 8.1-[1-(3-Azetidin-3-yl-4,5-dichloro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A solution of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl)ethyl]-5,6-dichloro-2-methoxyphenyl}azetidine-1-carboxylate(0.97 g, 1.9 mmol) in methylene chloride (20 mL) was treated withtrifluoroacetic acid (10 mL) and stirred at 20° C. for 30 min. Thereaction mixture was concentrated and the residue was diluted withmethanol (˜20 mL) and treated with saturated sodium bicarbonate solution(to pH˜8). The reaction mixture was concentrated to remove the methanol.The oil that was suspended in the aqueous layer was extracted into a 5:1mixture of dichloromethane/isopropanol, dried over magnesium sulfate,filtered, and concentrated to give the desired product (0.77 g, 99%)that was used in the next step without further purification. LCMS forC₁₈H₂₁Cl₂N₆O (M+H)⁺: m/z=407.1, 409.1; Found: 407.0, 409.0.

Step 9.1-{1-[4,5-Dichloro-3-(1-ethylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

A solution of1-[1-(3-azetidin-3-yl-4,5-dichloro-2-methoxyphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(40 mg, 0.098 mmol) in methanol (2.6 mL) was treated with sodiumcyanoborohydride (15 mg, 0.25 mmol) followed by acetaldehyde (22 μL,0.39 mmol) and stirred at 20° C. for 20 min. The reaction mixture wasquenched with acetic acid (130 μL, 2.3 mmol), diluted with methanol, andpurified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired product as a mixture ofenantiomers. This racemic mixture was separated by chiral HPLC (RT=18.6min and 22.0 min; Phenomenex Lux Cellulose C-4 column, 21.2×250 mm, 5micron particle size, eluting with 5% ethanol in hexanes at 18 ml/min,2.5 mg/inj) to give the desired peak 1 isomer (11 mg, 26%). ¹H NMR (300MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.45 (s, 1H), 7.33 (br s, 2H), 6.21 (q,J=6.9 Hz, 1H), 3.98-3.77 (m, 3H), 3.57 (s, 3H), 2.92-2.83 (m, 1H),2.79-2.72 (m, 1H), 2.55 (s, 3H), 2.35-2.22 (m, 2H), 1.70 (d, J=7.1 Hz,3H), 0.86 (t, J=7.1 Hz, 3H). LCMS for C₂₀H₂₅Cl₂N₆O (M+H)⁺: m/z=435.1;Found: 435.0.

Example 307.4-[1-(4-Amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-6-chloro-3-ethoxy-2-(1-isopropylazetidin-3-yl)benzonitrile

Step 1. tert-Butyl3-{3-[1-(4-amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidine-1-carboxylate

The desired compound was prepared according to the procedure of Example212, step 5 (chiral intermediate), using5-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine [ACES Pharma] instead of3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine as the starting material in18% yield. ¹H NMR (500 MHz, CDCl₃) δ 8.13 (s, 1H), 6.93 (br s, 1H), 6.79(s, 1H), 6.17 (q, J=7.1 Hz, 1H), 5.24 (s, 2H), 4.40-4.27 (m, 4H),4.27-4.18 (m, 1H), 4.03-3.92 (m, 1H), 3.80-3.70 (m, 1H), 2.43 (s, 3H),1.74 (d, J=7.1 Hz, 3H), 1.43 (s, 9H), 1.40 (t, J=7.0 Hz, 3H). LCMS forC₂₆H₃₂ClN₆O₃(M+H)⁺: m/z=511.2; Found: 511.2.

Step 2.4-[1-(4-Amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile

The desired compound was prepared according to the procedure of Example212, step 6, using tert-butyl3-{3-[1-(4-amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidine-1-carboxylateinstead of tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidine-1-carboxylateas the starting material in 99% yield. LCMS for C₂₁H₂₄ClN₆O (M+H)⁺:m/z=411.2; Found: 411.1.

Step 3.4-[1-(4-Amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-6-chloro-3-ethoxy-2-(1-isopropylazetidin-3-yl)benzonitrile

The desired compound was prepared according to the procedure of Example213 using4-[1-(4-amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrileinstead of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrileand acetone instead of formaldehyde as the starting materials in 65%yield. The product was isolated as a single enantiomer. ¹H NMR (300 MHz,dmso) δ 7.95 (s, 1H), 7.19 (s, 1H), 7.16-7.13 (m, 1H), 6.58 (s, 2H),6.11 (q, J=7.1 Hz, 1H), 4.04-3.67 (m, 5H), 3.04-2.92 (m, 2H), 2.36 (s,3H), 2.27-2.12 (m, 1H), 1.69 (d, J=7.1 Hz, 3H), 1.30 (t, J=6.9 Hz, 3H),0.85 (dd, J=6.1, 1.8 Hz, 6H). LCMS for C₂₄H₃₀ClN₆O (M+H)⁺: m/z=453.2;Found: 453.3.

Example 315.4-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}-3-methoxy-6-methylbenzonitrile

Step 1: 4-Acetyl-5-hydroxy-2-methylbenzonitrile

The 1-(4-bromo-2-hydroxy-5-methylphenyl)ethanone (8.5 g, 37 mmol, AlfaAesar catalog# H29125) was combined with zinc cyanide (8.7 g, 74 mmol)in N,N-dimethylformamide (75 mL) degassed with nitrogen and thetris(dibenzylideneacetone)dipalladium(0) (1.0 g, 1.1 mmol) and(9,9-dimethyl-9H-xanthene-4,5-diyl)bis(diphenylphosphine) (1.5 g, 2.6mmol) were added. The reaction was degassed again with nitrogen andheated to 120° C. and monitored by LC/MS. After heating for 18 h, thereaction was complete, the reaction was allowed to cool to roomtemperature, taken up in ethyl acetate and washed with water (2×),brine, dried over magnesium sulfate and concentrated to give the crudeproduct as a dark amber oil. The product was purified by FCC on silicagel eluting hexane: ethyl acetate gradient to give4-acetyl-5-hydroxy-2-methylbenzonitrile as a solid (6.3 g, 98%). LCMScalculated for C₁₀H₁₀NO₂ (M+H)⁺: m/z=176.1; found: 176.2.

Step 2: 4-Acetyl-3-hydroxy-2-iodo-6-methylbenzonitrile

The 4-acetyl-5-hydroxy-2-methylbenzonitrile (6.7 g, 38 mmol) wasdissolved in acetic acid (80 mL) and the N-Iodosuccinimide (10. g, 46mmol) was added. The reaction was heated to 80° C. in an oil bath andmonitored by LC/MS. After heating for 4 hrs the reaction was complete.This was allowed to cool and was concentrated in vacuo to give a darkoil. The oil was taken up in ethyl acetate and washed with water, sodiumbicarbonate (3×, until remained slightly basic), brine, dried overmagnesium sulfate and concentrated to give the crude product as a darkoil. The product was purified by FCC on silica gel eluting hexane: ethylacetate gradient to give 4-acetyl-3-hydroxy-2-iodo-6-methylbenzonitrileas pale yellow solid (7.2 g, 62%). LCMS calculated for C₁₀H₉INO₂ (M+H)⁺:m/z=301.9; found: 301.9.

Step 3: 4-Acetyl-2-iodo-3-methoxy-6-methylbenzonitrile

The 4-acetyl-3-hydroxy-2-iodo-6-methylbenzonitrile (5.0 g, 17 mmol) wasdissolved in N,N-dimethylformamide (50 mL) and the potassium carbonate(4.6 g, 33 mmol) and methyl iodide (2.1 mL, 33 mmol) were added. Thereaction was heated to 60° C. and monitored by LC/MS. After heating for2 hrs the reaction was complete. This was allowed to cool, diluted withethyl acetate (300 mL) and filtered to remove the remaining solids. Theorganic layer was washed with water (3×), brine, dried over magnesiumsulfate and concentrated to give the crude product as a dark solid. Theproduct was purified by FCC on silica gel eluting hexane: ethyl acetategradient to give 4-acetyl-3-methoxy-2-iodo-6-methylbenzonitrile as apale yellow crystalline solid (5.0 g, 96%). LCMS calculated forC₁₁H₁₁INO₂ (M+H)⁺: m/z=315.9; found: 316.0.

Step 4: tert-butyl3-(3-acetyl-6-cyano-2-methoxy-5-methylphenyl)azetidine-1-carboxylate

Zinc (1.70 g, 26.0 mmol) and celite (oven dried, 500 mg) were groundtogether in a flask until the solids appeared homogenous, the flask washeated with a heat gun while under high-vac for 5 minutes and thenback-filled with nitrogen. The solids were suspended inN,N-dimethylacetamide (4.2 mL) and 1,2-dibromoethane (0.13 mL, 1.5 mmol)was added. The reaction mixture was heated at 70° C. for 30 min and thencooled to room temperature. Chlorotrimethylsilane (0.16 mL, 1.3 mmol)was added dropwise and stirring was continued for 2 hrs at roomtemperature. A solution of tert-butyl 3-iodoazetidine-1-carboxylate(2.70 g, 9.52 mmol) in N,N-dimethylacetamide (4.35 mL) was then addedslowly and the resulting mixture was heated at 50° C. for 2 hrs. Thezinc-iodo reagent was allowed to cool to room temperature and was takenup in a syringe and filtered through a PTFE filter (adapted with aneedle) directly into a suspension oftris(dibenzylideneacetone)dipalladium(0) (0.111 g, 0.121 mmol) andtri-(2-furyl)phosphine (0.056 g, 0.24 mmol) and4-acetyl-2-iodo-3-methoxy-6-methylbenzonitrile (2.0 g, 6.3 mmol) inN,N-dimethylacetamide (19.6 mL) pre-degassed by bubbling N₂. Thereaction mixture was degassed with nitrogen again and heated to 70° C.After heating for 30 minutes the reaction was complete by LC/MS. Thiswas allowed to cool, taken up in ethyl acetate and washed with water,brine, dried over magnesium sulfate and concentrated to give the crudeproduct as an oil. The product was purified by FCC on silica gel elutinghexane; ethyl acetate gradient to give tert-butyl3-(3-acetyl-6-cyano-2-methoxy-5-methylphenyl)azetidine-1-carboxylate asa clear oil. (1.8 g, 82%). LCMS calculated for C₁₅H₁₇N₂O₄ (M+H)⁺:m/z=289.1; found: 289.1.

Step 5: tert-butyl3-[2-cyano-5-(1-hydroxyethyl)-6-methoxy-3-methylphenyl]azetidine-1-carboxylate

The tert-butyl3-(3-acetyl-6-cyano-2-methoxy-5-methylphenyl)azetidine-1-carboxylate(2.2 g, 6.4 mmol) was dissolved in methanol (20 mL) and cooled in icebath. The sodium tetrahydroborate (0.26 g, 7.0 mmol) was addedportionwise and the reaction was monitored by LC/MS. After stirring for1 h the reaction was complete. This was diluted with ethyl acetate andwater. The combined organic layer was washed with water, saturatedsodium bicarbonate, brine, dried over magnesium sulfate and concentratedto give crude tert-butyl3-[2-cyano-5-(1-hydroxyethyl)-6-methoxy-3-methylphenyl]azetidine-1-carboxylateas a yellow foam (2.1 g, 99%). LCMS calculated for C₁₅H₁₉N₂O₄ (M+H)⁺:m/z=291.1; found: 291.1.

Step 6: tert-butyl3-[3-(1-chloroethyl)-6-cyano-2-methoxy-5-methylphenyl]azetidine-1-carboxylate

The tert-butyl3-[2-cyano-5-(1-hydroxyethyl)-6-methoxy-3-methylphenyl]azetidine-1-carboxylate(2.1 g, 6.4 mmol) was taken up in methylene chloride (50.0 mL) andN,N-dimethylformamide (0.59 mL), cooled in an ice bath and the thionylchloride (0.56 mL, 7.7 mmol) was added slowly. After stirring for 2 hrsthe reaction was complete by LC/MS and was partitioned between ethylacetate and water. The combined organic layer was washed with watersaturated sodium bicarbonate, brine, dried over magnesium sulfate andconcentrated to give crude tert-butyl3-[3-(1-chloroethyl)-6-cyano-2-methoxy-5-methylphenyl]azetidine-1-carboxylateas an oil (2.2 g, 100%). LCMS calculated for C₁₅H₁₈ClN₂O₃(M+H)⁺:m/z=309.1; found: 309.1.

Step 7: tert-butyl3-{3-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-cyano-2-methoxy-5-methylphenyl}azetidine-1-carboxylate

The tert-butyl3-[3-(1-chloroethyl)-6-cyano-2-methoxy-5-methylphenyl]azetidine-1-carboxylate(2.3 g, 6.3 mmol) was dissolved in N,N-dimethylformamide (68 mL) withcesium carbonate (4.1 g, 13 mmol) and3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (1.4 g, 9.4 mmol) and washeated in an oil bath to 80° C. The reaction was stirred for 18 hrs andallowed to cool to room temperature. The reaction mixture was taken upin ethyl acetate, filtered, washed with water, brine, dried overmagnesium sulfate and concentrated to give the crude product. Theproduct was purified by FCC on silica gel eluting a (hexane: 10% ethanolethyl acetate) gradient to give tert-butyl3-{3-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-cyano-2-methoxy-5-methylphenyl}azetidine-1-carboxylateas a semisolid (1.5 g, 50%). LCMS calculated for C₂₅H₃₂N₇O₃ (M+H)⁺:m/z=478.2; found: 478.2. The enantiomers were separated by Chiral columnHPLC using: Phenomenex LUX Cellulose Column, 21.1×250 mm, 5 micron, 15%ethanol in hexane, 18 mL/min˜5 mg/injection to give: First peakretention time: 2.1 minutes, tert-butyl3-{3-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-cyano-2-methoxy-5-methylphenyl}azetidine-1-carboxylate;Second peak retention time: 3.9 minutes, tert-butyl3-{3-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-cyano-2-methoxy-5-methylphenyl}azetidine-1-carboxylate.

Step 8:4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-methoxy-6-methylbenzonitrilebis(trifluoroacetate)

The tert-butyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-cyano-2-methoxy-5-methylphenyl}azetidine-1-carboxylate(0.35 g, 0.73 mmol) (Step 7, peak 1) was dissolved in methylene chloride(3.0 mL) and trifluoroacetic acid (1.0 mL) at room temperature. Afterstirring for 1 h the reaction was complete by LC/MS. The reaction wasconcentrated in vacuo to give4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-methoxy-6-methylbenzonitrile(bis(trifluoroacetate) as a viscous amber oil (0.50 g, 100%). LCMScalculated for C₂₀H₂₄N₇O (M+H)⁺: m/z=378.2; found: 378.2.

Step 9:4-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}-3-methoxy-6-methylbenzonitrile

The4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-methoxy-6-methylbenzonitrilebis(trifluoroacetate) (0.074 g, 0.10 mmol) was dissolved in ethanol (3.0mL) and DIPEA (0.071 mL, 0.41 mmol) and the (S)-(−)-methyloxirane(0.0071 g, 0.12 mmol) was added. The reaction was heated in a sealedtube to 90° C. and monitored by LC/MS. After heating for 6 hrs thereaction was purified without workup by prep HPLC on a C-18 columneluting water: acetonitrile gradient buffered pH 10 to give the titlecompound as a white amorphous solid (0.018 g, 40%). The product wasisolated as a single enantiomer. LCMS calculated for C₂₃H₃₀N₇O₂ (M+H)⁺:m/z=436.2; found: 436.3. ¹H NMR (300 MHz, DMSO-d₆) δ 8.09 (s, 1H), 7.21(s, 1H), 6.22 (q, J=7.1 Hz, 1H), 4.34 (d, J=4.5 Hz, 1H), 4.09-3.83 (m,3H), 3.60 (s, 3H), 3.58-3.51 (m, 1H), 3.12-2.95 (m, 2H), 2.55 (s, 3H),2.33 (s, 3H), 2.27 (d, J=5.9 Hz, 2H), 1.71 (d, J=7.1 Hz, 3H), 1.00 (d,J=6.2 Hz, 3H).

Example 316.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[6-(1-hydroxy-1-methylethyl)pyridin-3-yl]benzonitrile

Step 1. 5-bromo-N-methoxy-N-methylpyridine-2-carboxamide

N, O-dimethylhydroxylamine hydrochloride (500 mg, 5 mmol) was added to amixture of N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uroniumhexafluorophosphate (1400 mg, 3.7 mmol), N,N-diisopropylethylamine (1000μL, 7 mmol) and 5-bromopyridine-2-carboxylic acid (500 mg, 2 mmol,Frontier Scientific catalog# B1704) in N,N-dimethylformamide (10 mL).The reaction mixture was stirred overnight at room temperature and wascomplete by LC/MS. The reaction was partitioned between water and EtOAc.The combined organic layer was washed with brine, dried over MgSO₄,filtered and concentrated to give the crude product. The product waspurified on by FCC on silica gel eluting a hexane: EtOAc (0-30%)gradient to give 5-bromo-N-methoxy-N-methylpyridine-2-carboxamide clearoil (0.50 g, 60%). LCMS calculated for C₈H₁₀BrN₂O₂ (M+H)⁺: m/z=244.9,246.9; found: 244.9, 246.9.

Step 2. 1-(5-bromopyridin-2-yl)ethanone

Methylmagnesium chloride 3.0 M in THF (0.5 mL) was added dropwise to amixture of 5-bromo-N-methoxy-N-methylpyridine-2-carboxamide (200 mg, 0.8mmol) in tetrahydrofuran (10 mL) at 0° C. After stirring for 1 hr atroom temperature, the reaction was quenched with 1 N NH₄Cl and wasextracted with EtOAc. The combined organic layer was washed with brineand dried over MgSO₄, concentrated to give the crude product1-(5-bromopyridin-2-yl)ethanone (0.15 g, 90%). LCMS calculated forC₇H₇BrNO (M+H)⁺: m/z=199.9, 201.9; found: 199.9, 201.9.

Step 3. 2-(5-bromopyridin-2-yl)propan-2-ol

Methylmagnesium chloride 3.0 M in THF (0.3 mL) was added dropwise to amixture of 1-(5-bromopyridin-2-yl)ethanone (100 mg, 0.5 mmol) intetrahydrofuran (10 mL) at 0° C. After stirring for 1 h at roomtemperature, the reaction was quenched with 1 N NH₄Cl and was extractedwith EtOAc. The combined organic layer was washed with brine and driedover MgSO₄, concentrated to give crude2-(5-bromopyridin-2-yl)propan-2-ol (0.1 g, 100%). LCMS calculated forC₈H₁₁BrNO (M+H)⁺: m/z=215.9, 217.9; found: 215.8, 217.8.

Step 4. [6-(1-hydroxy-1-methylethyl)pyridin-3-yl]boronic acid

A mixture of 2-(5-bromopyridin-2-yl)propan-2-ol (70 mg, 0.3 mmol),4,4,5,5,4′,4′,5′,5′-octamethyl-[2,2′]bi[[1,3,2]dioxaborolanyl] (90. mg,0.36 mmol), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II),complex with dichloromethane (1:1) (10 mg, 0.01 mmol), and potassiumacetate (100 mg, 1 mmol) in 1,4-dioxane (5 mL) was heated at 120° C.overnight. The reaction was complete by LC/MS, was concentrated in vacuoto give crude [6-(1-hydroxy-1-methylethyl)pyridin-3-yl]boronic acid.LCMS calculated for CsH₁₃BNO₃ (M+H)⁺: m/z=182.1; found: 182.1.

Step 5.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[6-(1-hydroxy-1-methylethyl)pyridin-3-yl]benzonitrilebis(2,2,2-trifluoroacetate)

Sodium carbonate (10 mg, 0.09 mmol) in water (0.5 mL) was added to amixture of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-bromo-6-chloro-3-ethoxybenzonitrile(20 mg, 0.04 mmol, racemic intermediate from Example 43, Step 5) and[6-(1-hydroxy-1-methylethyl)pyridin-3-yl]boronic acid (12 mg, 0.069mmol, Example 306, Step 4) in acetonitrile (1 mL). The reaction mixturewas degassed with N₂ and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (2 mg, 0.002 mmol) was added. The reactionwas degassed with N₂ again and heated to 100° C. for 1 h. The reactionwas allowed to cool to room temperature and was purified without workupby prep HPLC on a C-18 column eluting a water; acetonitrile gradientbuffered with TFA to give the title compound as white amorphous solid.The product was isolated as a racemic mixture. LCMS calculated forC₂₅H₂₇ClN₇O₂(M+H)⁺: m/z=492.1; found: 492.1. ¹H NMR (500 MHz, DMSO-d₆) δ8.60 (d, J=2.0 Hz, 1H), 8.22 (s, 1H), 7.96 (dd, J=8.2, 2.3 Hz, 1H), 7.80(d, J=8.3 Hz, 1H), 7.73 (s, 1H), 6.36 (q, J=7.0 Hz, 1H), 3.52-3.40 (m,1H), 3.40-3.30 (m, 1H), 2.59 (s, 3H), 1.80 (d, J=7.0 Hz, 3H), 1.48 (d,J=2.3 Hz, 6H), 0.88 (t, J=7.0 Hz, 3H).

Example 318.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile

Step 1. 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile

The 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile was prepared byanalogous methods described in Example 43, Step 1 and Step 2, but usingN-iodosuccinimide. LCMS calculated for C₁₁H₁₀ClINO₂ (M+H)⁺: m/z=349.9;found: 350.0

Step 2. 4-acetyl-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile

The 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile (0.20 g, 0.57 mmol)was combined with pyrrolidine (0.052 mL, 0.63 mmol) inN,N-dimethylformamide (2.0 mL) with cesium carbonate (0.19 g, 0.57 mmol)and heated to 120° C. in a sealed tube. After heating for 18 hrs thereaction was allowed to cool, taken up in ethyl acetate, washed withwater, brine, dried over magnesium sulfate and concentrated to give thecrude product as a dark oil. The product was purified by FCC on silicagel eluting with hexane: ethyl acetate gradient to give4-acetyl-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile as an oil(0.045 g, 27%). LCMS calculated for C₁₅H₁₈ClN₂O₂(M+H)⁺: m/z=293.1; found293.1.

Step 3.6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-pyrrolidin-1-ylbenzonitrile

The 4-acetyl-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile (0.045 g,0.15 mmol) was dissolved in methanol (3 mL) and cooled in an ice bath.The sodium tetrahydroborate (0.0058 g, 0.15 mmol) was added and thereaction was monitored by LC/MS. After stirring for 1 h, the reactionwas taken up in ethyl acetate and washed with water, sodium bicarbonate,brine and dried over magnesium sulfate to give crude6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-pyrrolidin-1-ylbenzonitrile as aclear oil (0.045 g, 100%). LCMS calculated for C₁₅H₂₀ClN₂O₂(M+H)⁺:m/z=295.1; found 295.1.

Step 4.6-chloro-4-(1-chloroethyl)-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile

The 6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-pyrrolidin-1-ylbenzonitrile(0.045 g, 0.15 mmol) was taken up in methylene chloride (3.0 mL) andN,N-dimethylformamide (0.002 mL, 0.03 mmol) and cooled in an ice bath.The thionyl chloride (0.017 mL, 0.23 mmol) was added and the reactionwas monitored by LC/MS. After stirring for 2 hrs the reaction wascomplete. The reaction was then taken up in ethyl acetate, washed withsodium bicarbonate, brine, dried over magnesium sulfate and concentratedto give crude6-chloro-4-(1-chloroethyl)-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile as ayellow oil (0.048 g, 100%). LCMS calculated for C₁₅H₁₉Cl₂N₂O (M+H)⁺:m/z=313.1; found 313.1.

Step 5.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile

The 6-chloro-4-(1-chloroethyl)-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile(0.048 g, 0.15 mmol, racemic mixture) was combined with3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.034 g, 0.23 mmol) andcesium carbonate (0.10 g, 0.31 mmol) in N,N-dimethylformamide (3.0 mL)and heated in an oil bath to 85° C. After heating for 18 hrs thereaction was complete. The crude reaction was purified with out work upby prep HPLC on a C-18 column eluting water: acetonitrile gradientbuffered pH 10 to give the title compound as a white amorphous solid(0.012 g, 18%). The product was isolated as a racemic mixture. LCMScalculated for C₂₁H₂₅ClN₇O (M+H)⁺: m/z=426.1; found 426.1. ¹H NMR (300MHz, DMSO-d₆) δ 8.11 (s, 1H), 6.91 (s, 1H), 6.25 (q, J=7.1 Hz, 1H), 3.71(dp, J=15.7, 8.1, 7.2 Hz, 4H), 3.49-3.35 (m, 2H), 2.55 (s, 3H),2.00-1.76 (m, 4H), 1.70 (d, J=7.1 Hz, 3H), 1.34 (t, J=7.0 Hz, 3H).

Example 319.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile

Step 1.4-acetyl-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile

To a mixture of 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile (50 mg,0.1 mmol, Example 318, Step 1), 3-methoxyazetidine hydrochloride (21 mg,0.17 mmol Chem-Impex catalog#20140) and cesium carbonate (70. mg, 0.21mmol) in 1,4-dioxane (4 mL) was added(9,9-dimethyl-9H-xanthene-4,5-diyl)bis(diphenylphosphine) (40 mg, 0.07mmol) and tris(dibenzylideneacetone)dipalladium (0) (60 mg, 0.07 mmol).The reaction mixture was degassed with N₂. The reaction was heated at80° C. for 2 hrs and was monitored by LC/MS. The reaction was allowed tocool to room temperature, was diluted with water and extracted withEtOAc. The combined organic layers were washed with brine, dried overMgSO4, filtered and concentrated to give the crude product. The productwas purified by FCC on silica gel eluting (hexanes: EtOAc 0-70%)gradient to give to4-acetyl-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile asclear oil (0.030 g, 70%). LCMS calculated for C₁₅H₁₈ClN₂O₃(M+H)⁺:m/z=309.1; found: 309.1.

Step 2.6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(3-methoxyazetidin-1-yl)benzonitrile

4-Acetyl-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile (30mg, 0.1 mmol was dissolved in methanol (5 mL) cooled to 0° C. and sodiumtetrahydroborate (5.5 mg, 0.14 mmol) was added. Reaction was stirred for1 h at 0° C. The reaction was partitioned between EtOAc and water. Thecombined organic layer was washed with water and saturated NaHCO₃,brine, dried over Na₂SO₄, filtered and concentrated to give crude6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(3-methoxyazetidin-1-yl)benzonitrile(0.030 g, 100%). LCMS calculated for C₁₅H₂₀ClN₂O₃ (M+H)⁺: m/z=311.1;found: 311.1.

Step 3.6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile

6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(3-methoxyazetidin-1-yl)benzonitrile(30 mg, 0.1 mmol) (racemic mixture) was dissolved in methylene chloride(5 mL) and N,N-dimethylformamide (100 μL, 1 mmol). Thionyl chloride (18μL, 0.24 mmol) was added dropwise at room temperature and the reactionwas stirred for 2 hrs. The reaction was diluted with EtOAc, washed withwater and saturated NaHCO₃, brine, dried over Na₂SO₄, filtered andconcentrated to give the crude6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile(0.030 g, 100%). LCMS calculated for C₁₅H₁₉Cl₂N₂O₃ (M+H)⁺: m/z=329.1;found: 329.1.

Step 4.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile

Cesium carbonate (50 mg, 0.2 mmol) was added to a mixture of3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (16 mg, 0.10 mmol) and6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile(30 mg, 0.09 mmol) in N,N-dimethylformamide (3 mL, 40 mmol) and thereaction was stirred at 80° C. overnight. The mixture was diluted withEtOAc, washed with water, brine, dried over Na₂SO₄, filtered andconcentrated the crude product. The product was purified was purified byprep HPLC on a C-18 column eluting water: acetonitrile gradient bufferedpH 10 to give the title compound as a white amorphous solid (0.007 g,20%). The product was isolated as a racemic mixture. LCMS calculated forC₂₁H₂₅ClN₇O₂(M+H)⁺: m/z=442.1; found: 442.1. H NMR (400 MHz, DMSO-d₆) δ8.11 (s, 1H), 6.80 (s, 1H), 6.18 (d, J=7.1 Hz, 1H), 4.58-4.44 (m, 2H),4.18 (m, 1H), 4.13-4.01 (m, 2H), 3.81-3.62 (m, 2H), 3.23 (s, 3H), 2.55(s, 3H), 1.69 (d, J=7.1 Hz, 3H), 1.35 (t, J=7.0 Hz, 3H).

Example 320.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-(1-isopropylazetidin-3-yl)-6-methylbenzonitrile

Step 1:4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-ethoxy-6-methylbenzonitrilebis(trifluoroacetate)

Using methods described in Example 315 but using ethyl iodide in Step 3,instead of methyl iodide, the intermediate4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-ethoxy-6-methylbenzonitrilebis(trifluoroacetate) was prepared. LCMS calculated for C₂₁H₂₆N₇O(M+H)⁺: m/z=392.2; found: 392.2.

Step 2.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-(1-isopropylazetidin-3-yl)-6-methylbenzonitrile

To a mixture of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-ethoxy-6-methylbenzonitrile(70 mg, 0.2 mmol) in methanol (50 mL) was added acetone (0.1 mL, 2 mmol)and sodium cyanoborohydride (17 mg, 0.27 mmol). The reaction was stirredat room temperature for 1 h, and was complete by LC/MS. The reaction wasquenched with water and was extracted with EtOAc. The combined organiclayer was washed with brine, dried over MgSO₄, filtered and concentratedto give the crude product. The product was purified by prep HPLC on aC-18 column eluting water: acetonitrile gradient buffered pH 10 to givethe title compound as a white amorphous solid (0.030 g, 40%). Theproduct was isolated as a racemic mixture. LCMS calculated for C₂₄H₃₂N₇O(M+H)⁺: m/z=434.2; found: 434.3. ¹H NMR (300 MHz, CD₃OD) δ 8.17 (s, 1H),7.35 (s, 1H), 6.37 (q, J=7.1 Hz, 1H), 4.17-3.98 (m, 4H), 3.90-3.71 (m,3H), 2.65 (s, 3H), 2.46 (s, 4H), 1.84 (d, J=7.1 Hz, 3H), 1.42 (t, J=7.0Hz, 3H), 1.03 (dd, J=6.2, 1.4 Hz, 6H).

Example 321.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-[1-(2-hydroxy-2-methylpropyl)azetidin-3-yl]-6-methylbenzonitrile

The4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-ethoxy-6-methylbenzonitrile(0.055 g, 0.14 mmol, chiral intermediate from Example 320, Step 1) wascombined with tetrahydrofuran (22 mL), DIPEA (0.049 mL, 0.28 mmol) andoxirane, 2,2-dimethyl- (0.018 mL, 0.21 mmol) at room temperature. Thereaction was heated to 95° C. and allowed to stir overnight. Thereaction was allowed to cool to room temperature and was purifiedwithout workup by prep HPLC on a C-18 column eluting water: acetonitrilegradient buffered pH 10 to give the title compound as a white amorphoussolid (0.035 g, 50%). The product was isolated as a single enantiomer.LCMS calculated for C₂₅H₃₄N₇O₂ (M+H)⁺: m/z=464.3; found: 464.3. ¹H NMR(300 MHz, DMSO-d₆) δ 8.09 (s, 1H), 7.23 (s, 1H), 6.21 (q, J=6.8 Hz, 1H),4.00 (m, 4H), 3.81-3.54 (m, 2H), 3.15 (m, 2H), 2.53 (s, 3H), 2.33 (s,3H), 2.27 (bs, 2H), 1.70 (d, J=7.1 Hz, 3H), 1.30 (t, J=6.9 Hz, 3H), 1.04(s, 6H).

Example 322.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-[1-(2-hydroxy-2-methylpropanoyl)azetidin-3-yl]-6-methylbenzonitrile

The4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-3-ethoxy-6-methylbenzonitrile(0.075 g, 0.10 mmol, chiral intermediate from Example 320, Step 1) wasdissolved in N,N-dimethylformamide (3.0 mL) and DIPEA (0.089 mL, 0.51mmol) and the propanoic acid, 2-hydroxy-2-methyl- (0.013 g, 0.12 mmol)and N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uroniumhexafluorophosphate (0.058 g, 0.15 mmol) were added. The reaction wasstirred at room temperature for 18 hrs and was complete by LC/MS. Theproduct was purified without workup by prep HPLC on a C-18 columneluting water: acetonitrile gradient buffered to pH 10 to give the titlecompound as a white amorphous solid (0.025 g, 51%). The product wasisolated as a single enantiomer. LCMS calculated for C₂₅H₃₂N₇O₃ (M+H)⁺:m/z=478.2; found: 478.2. ¹H NMR (300 MHz, DMSO-d₆) δ 8.10 (s, 1H), 7.29(s, 1H), 6.24 (q, J=6.8 Hz, 1H), 5.07 (s, 1H), 4.90-4.75 (m, 1H),4.73-4.58 (m, 1H), 4.39 (p, J=8.5 Hz, 1H), 4.30-4.05 (m, 2H), 3.75 (d,J=7.1 Hz, 2H), 2.54 (s, 3H), 2.38 (s, 3H), 1.72 (d, J=6.9 Hz, 3H), 1.35(t, J=6.1 Hz, 3H), 1.26 (s, 3H), 1.23 (s, 3H).

Examples 310 and 311. Diastereoisomers of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}pyrrolidin-2-one

Step 1. 1-(5-Chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanol

The desired compound was prepared according to the procedure of Example212, step 4 (racemic mixture), using1-(5-chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanone instead oftert-butyl3-(3-acetyl-5-chloro-6-cyano-2-ethoxyphenyl)azetidine-1-carboxylate asthe starting material in 94% yield as a 96:4 mixture of enantiomers(RT=3.56 min and 4.28 min; Chiral Technologies ChiralPak AD-H column,20×250 mm, 5 micron particle size, eluting with 5% ethanol in hexanes at1 ml/min). LCMS for C₁₁H₁₃ClIO (M-(OH))⁺: m/z=323.0; Found: 322.9.

Step 2.1-[1-(5-Chloro-2-ethoxy-3-iodo-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine

The desired compound was prepared according to the procedure of Example212, step 5, using 1-(5-Chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanol(96:4 mixture from step 1) instead of tert-butyl3-[3-chloro-2-cyano-6-ethoxy-5-(1-hydroxyethyl)phenyl]azetidine-1-carboxylateas the starting material in 32% yield as a single enantiomer (peak 1desired, retention time=3.39 min; ChiralPak IA column, 20×250 mm, 5micron particle size, eluting with 3% ethanol in hexanes at 18 ml/min).LCMS for C₁₇H₂₀ClIN₅O (M+H)⁺: m/z=472.0; Found: 472.0.

Step 3. Methyl(2E)-3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}acrylate

A suspension of1-[1-(5-chloro-2-ethoxy-3-iodo-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine(peak 1 single isomer from step 2) (0.61 g, 1.3 mmol) in acetonitrile(7.4 mL) in a sealed tube was degassed with nitrogen and treated withtriphenylphosphine (0.048 g, 0.18 mmol), methyl acrylate (0.41 mL, 4.5mmol), and palladium acetate (0.029 g, 0.13 mmol) followed bytriethylamine (0.54 mL, 3.9 mmol) and heated at 100° C. for 16 h. Thereaction mixture was cooled to room temperature, filtered, and thesolids washed with acetonitrile. The filtrate was concentrated to aresidue. The crude material was purified by flash column chromatographyusing ethyl acetate (containing 3% methanol) in hexanes (0%-100%) togive the desired product (0.40 g, 72%). LCMS for C₂₁H₂₅ClN₅O₃ (M+H)⁺:m/z=430.2; Found: 430.2.

Step 4. Diastereoisomers of methyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-4-nitrobutanoate

A solution of methyl(2E)-3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}acrylate(0.40 g, 0.93 mmol) in nitromethane (6.3 mL) was treated with1,8-diazabicyclo[5.4.0]undec-7-ene (0.14 mL, 0.93 mmol) and stirred at90° C. for 22 h. The reaction mixture was concentrated, diluted withmethanol, and purified by preparative LCMS (XBridge C18 Column, elutingwith a gradient of acetonitrile in water with 0.1% trifluoroacetic acid,at flow rate of 60 mL/min). The LCMS fractions were concentrated toremove acetonitrile, treated with solid sodium bicarbonate, andextracted into ethyl acetate. The ethyl acetate was concentrated to givethe desired product (0.22 g, 48%) as a mixture of diastereoisomers. LCMSfor C₂₂H₂₈ClN₆O₅(M+H)⁺: m/z=491.2; Found: 491.2.

Step 5. Diastereoisomers of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}pyrrolidin-2-on

A solution of methyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-4-nitrobutanoate(0.089 g, 0.18 mmol) in methanol (1.3 mL) was treated with nickelchloride hexahydrate (0.087 g, 0.36 mmol) was and stirred for 5 min. Thereaction mixture was cooled to 0° C., treated with sodiumtetrahydroborate (0.073 g, 1.9 mmol) in four portions, and stirred atroom temperature for 30 min. The reaction mixture was heated at 60° C.for 1.5 h, cooled to room temperature, diluted with saturated sodiumbicarbonate solution (10 mL) and dichloromethane (25 mL), and filteredthrough Celite. The Celite was washed with dichloromethane and thefiltrate was transferred to a separatory funnel. The organic layer wasseparated, washed with brine, dried over sodium sulfate, filtered, andconcentrated to residue. The crude residue was diluted with methanol andpurified by preparative LCMS (XBridge C18 column, eluting with agradient of acetonitrile/water containing 0.1% ammonium hydroxide, atflow rate of 60 mL/min) to give the desired peak 1 diastereoisomer (16mg, 21%) and peak 2 diastereoisomer (19 mg, 24%). Peak 1 (compound 310):¹H NMR (300 MHz, DMSO-d₆) δ 8.10 (s, 1H), 7.89 (s, 1H), 7.34 (s, 1H),6.21 (q, J=7.1 Hz, 1H), 4.38-4.22 (m, 1H), 3.93-3.80 (m, 1H), 3.79-3.67(m, 1H), 3.65-3.55 (m, 1H), 3.28-3.20 (m, 1H), 2.54 (s, 3H), 2.29 (dd,J=17.5, 8.3 Hz, 1H), 2.21 (s, 3H), 1.70 (d, J=7.0 Hz, 3H), 1.40 (t,J=6.9 Hz, 3H). LCMS for C₂₁H₂₆ClN₆O₂(M+H)⁺: m/z=429.2; Found: 429.2.Peak 2 (compound 311): H NMR (300 MHz, DMSO-d₆) δ 8.11 (s, 1H), 7.89 (s,1H), 7.33 (s, 1H), 6.20 (q, J=7.1 Hz, 1H), 4.38-4.22 (m, 1H), 3.90-3.68(m, 2H), 3.65-3.56 (m, 1H), 3.28-3.17 (m, 1H), 2.54 (s, 3H), 2.32 (dd,J=17.3, 8.5 Hz, 1H), 2.21 (s, 3H), 1.69 (d, J=7.0 Hz, 3H), 1.39 (t,J=6.9 Hz, 3H). LCMS for C₂₁H₂₆ClN₆O₂(M+H)⁺: m/z=429.2; Found: 429.2.

Example 323.4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile

Step 1. 4-Acetyl-6-chloro-3-ethoxy-2-vinylbenzonitrile

A mixture of 4-acetyl-6-chloro-3-ethoxy-2-iodobenzonitrile (1.3 g, 3.6mmol), 4,4,5,5-tetramethyl-2-vinyl-1,3,2-dioxaborolane (740 μL, 4.3mmol), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II),complex with dichloromethane (1:1) (100 mg, 0.20 mmol) and potassiumcarbonate (1.5 g, 11 mmol) in 1,4-dioxane (20 mL) and water (10 mL) washeated at 80° C. overnight. The mixture was cooled to room temperatureand extracted with ethyl acetate. The extracts were washed with brine,dried over sodium sulfate, filtered and concentrated. Purification onsilica gel using ethyl acetate in hexanes (0-20%) gave the desiredcompound, 780 mg, 87%. LCMS calculated for Cl₃H₁₃ClNO₂ (M+H)⁺:m/z=250.1; found: 250.1. ¹H NMR (400 MHz, DMSO-d₆): δ 7.78 (s, 1H), 6.83(m, 1H), 6.10 (m, 1H), 5.83 (m, 1H), 3.84 (m, 2H), 2.58 (s, 3H), 1.22(m, 3H).

Step 2. tert-Butyl[2-(3-acetyl-5-chloro-6-cyano-2-ethoxyphenyl)-2-hydroxyethyl]carbamate

0.2 M Osmium tetraoxide in water (0.5 mL) was added to a solution oftert-butyl [(4-chlorobenzoyl)oxy]carbamate (Ref. Lawrence Harris, J.Org. Chem, 2011, 76, 358-372). (0.91 g, 3.3 mmol) in acetonitrile (10mL) and stirred for 10 minutes.4-Acetyl-6-chloro-3-ethoxy-2-vinylbenzonitrile (0.56 g, 2.2 mmol) as asolution in acetonitrile (10 mL) was added to the carbamate solutionfollowed by the addition of water (2 mL) and the reaction was stirredfor 3 hours at room temperature. The reaction was quenched withsaturated 10 M dipotassium disulfite in water (12 mL) and stirred for 5minutes. Water was added and the reaction mixture was extracted withethyl acetate. The extracts were washed with saturated sodiumbicarbonate solution, brine and dried over sodium sulfate, filtered andevaporated. Purification on silica gel using ethyl acetate in hexane(0-100%) gave the desired compound as a racemic mixture, 610 mg, 72%.LCMS calculated for C₁₈H₂₄ClN₂O₅(M+H)⁺: m/z=383.1; found: 383.1. ¹H NMR(400 MHz, DMSO-d₆): δ 7.62 (s, 1H), 7.03 (br s, 1H), 5.68 (br s, 1H),3.96 (m, 1H), 3.69 (m, 1H), 3.31 (m, 1H), 3.19 (m, 1H), 2.60 (s, 3H),1.30 (m, 12H).

Step 3.4-Acetyl-6-chloro-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile

tert-Butyl[2-(3-acetyl-5-chloro-6-cyano-2-ethoxyphenyl)-2-hydroxyethyl]carbamate(290 mg, 0.76 mmol) (racemic mixture from step 2) was treated with 4.0 Mhydrogen chloride in 1,4-dioxane (6.1 mL) for 15 minutes and the mixturewas evaporated. The residue was dissolved in tetrahydrofuran (2.3 mL)and N,N-diisopropylethylamine (0.66 mL, 3.8 mmol).N,N-carbonyldiimidazole (250 mg, 1.5 mmol) was added and the reactionmixture was refluxed at 70° C. overnight. The reaction mixture wasevaporated. Purification on silica gel using ethyl acetate in hexane(0-100%) gave the desired compound as a racemic mixture, 110 mg, 47%.LCMS calculated for C₁₄H₁₄ClN₂O₄(M+H)⁺: m/z=309.1; found: 309.1. ¹H NMR(400 MHz, DMSO-d₆): δ 8.00 (br s, 1H), 7.93 (s, 1H), 5.99 (m, 1H), 3.89(m, 1H), 3.81 (m, 2H), 3.52 (m, 1H), 2.58 (s, 3H), 1.23 (m, 3H).

Step 4.6-Chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile

Sodium tetrahydroborate (19 mg, 0.50 mmol) was added to a mixture of4-acetyl-6-chloro-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile(100 mg, 0.34 mmol) (racemic mixture from step 3) in methanol (1.6 mL,38 mmol) at 0° C. and the reaction mixture was stirred at roomtemperature for 10 minutes and evaporated. The residue was diluted withethyl acetate, washed with 1 N HCl, brine, dried over sodium sulfate,filtered and concentrated to give the desired compound as a mixture offour diastereomers, 58 mg, 55%. LCMS calculated for C₁₄H₁₆ClN₂O₄(M+H)⁺:m/z=311.1; found: 311.1.

Step 5.6-Chloro-4-(1-chloroethyl)-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile

To a mixture of6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile(58 mg, 0.19 mmol) (mixture of four diastereomers from step 4),N,N-dimethylformamide (36 μL) in methylene chloride (1 mL), thionylchloride (40 μL, 0.56 mmol) was added and the mixture was stirred atroom temperature for 20 minutes The mixture was diluted with methylenechloride, washed with saturated sodium bicarbonate, water, brine, driedover sodium sulfate, filtered and concentrated to give the desiredcompound as a mixture of four diastereomers, 55 mg, 91%. LCMS calculatedfor C₁₄H₁₅Cl₂N₂O₃ (M+H)⁺: m/z=329.0; found: 329.1.

Step 6.4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile

Cesium Carbonate (0.11 g, 0.34 mmol) was added to a mixture of3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (30 mg, 0.20 mmol) (mixtureof four diastereomers from step 5) in N,N-dimethylformamide (0.91 mL)and stirred for 10 minutes. To the mixture was added6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile(56 mg, 0.17 mmol) in N,N-dimethylformamide (1.0 mL) and the reactionwas stirred at 90° C. for 1 hour. Purification by preparative LCMS (pH10) using RP-HPLC (XBridge C18 column, eluting with a gradient ofacetonitrile/water containing 0.1% ammonium hydroxide, at flow rate of30 mL/min) gave the desired compounds as Peak 1 (racemic mixture of twodiastereomers) LCMS calculated for C₂₀H₂₁ClN₇O₃(M+H)⁺: m/z=442.1; found:442.1. ¹H NMR (400 MHz, DMSO-d₆): δ 8.17 (s, 1H), 8.00 (br s, 1H), 7.79(s, 1H), 6.25 (m, 1H), 5.92 (m, 1H), 3.90 (m, 3H), 3.57 (m, 1H), 2.58(s, 3H), 1.75 (m, 3H), 1.40 (m, 3H); Peak 2 (racemic mixture of 2diastereomers): LCMS calculated for C₂₀H₂₁ClN₇O₃(M+H)⁺: m/z=442.1;found: 442.1. ¹H NMR (400 MHz, DMSO-d₆): δ 8.12 (s, 1H), 8.00 (br s,1H), 7.71 (s, 1H), 6.23 (m, 1H), 5.96 (m, 1H), 3.85 (m, 3H), 3.58 (m,1H), 2.58 (s, 3H), 1.75 (m, 3H), 1.40 (m, 3H).

Chiral purification of Peak 2 (racemic mixture of two diastereomers) onPhenomenex Lux Cellulose-1, 21.2×250 mm, 5 micron particle size at 18mL/min using 20% ethanol in hexanes gave Peak 3 and Peak 4. Peak 3,retention time=12.22 minutes (single enantiomer): LCMS calculated forC₂₀H₂₁ClN₇O₃(M+H)⁺: m/z=442.1; found: 442.1. ¹H NMR (400 MHz, DMSO-d₆):δ 8.12 (s, 1H), 7.98 (br s, 1H), 7.71 (s, 1H), 6.23 (m, 1H), 5.96 (m,1H), 3.85 (m, 3H), 3.58 (m, 1H), 2.58 (s, 3H), 1.75 (m, 3H), 1.40 (m,3H). Peak 4, retention time=16.25 minutes (single enantiomer). LCMScalculated for C₂₀H₂₁ClN₇O₃(M+H)⁺: m/z=442.1; found: 442.1. ¹H NMR (400MHz, DMSO-d₆): δ 8.12 (s, 1H), 7.98 (br s, 1H), 7.71 (s, 1H), 6.23 (m,1H), 5.96 (m, 1H), 3.85 (m, 3H), 3.58 (m, 1H), 2.58 (s, 3H), 1.75 (m,3H), 1.40 (m, 3H).

Example 324.6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}morpholin-3-one

Step 1. 1-(5-Chloro-2-methoxy-4-methyl-3-vinylphenyl)ethanone

A mixture of 1-(3-bromo-5-chloro-2-methoxy-4-methylphenyl)ethanone (2.6g, 9.5 mmol), 4,4,5,5-tetramethyl-2-vinyl-1,3,2-dioxaborolane (1.9 mL,11 mmol), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II),complex with dichloromethane (1:1) (400 mg, 0.5 mmol) and potassiumcarbonate (4.0 g, 29 mmol) in 1,4-dioxane (60 mL), and water (30 mL).The resulting mixture was heated at 80° C. for 3 hours. The mixture wascooled to room temperature and extracted with ethyl acetate.Purification on a silica gel using ethyl acetate in hexanes (0-20%) gavethe desired compound, 2.0 g, 94%. LCMS calculated for C₁₂H₁₄ClO₂ (M+H)⁺:m/z=225.1; found: 225.1.

Step 2. tert-Butyl[2-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-2-hydroxyethyl]carbamate

0.2 M Osmium tetraoxide in water (1 mL) was added to a solution oftert-butyl [(4-chlorobenzoyl)oxy]carbamate (2.0 g, 7.2 mmol) (Ref.Lawrence Harris, J. Org. Chem, 2011, 76, 358-372) in acetonitrile (22mL) and stirred for 10 minutes.1-(5-Chloro-2-methoxy-4-methyl-3-vinylphenyl)ethanone (1.1 g, 4.8 mmol)as a solution in acetonitrile (22 mL) was added to the carbamatesolution followed by the addition of water (5 mL). The reaction wasstirred for 3 hours at room temperature. The reaction was quenched withsaturated 10 M dipotassium disulfite in water (25 mL) and stirred for 5minutes. Water was added to the reaction and the mixture was extractedwith ethyl acetate. The organic extracts were washed with saturatedsodium bicarbonate solution, brine, dried over sodium sulfate andevaporated under reduced pressure. Purification on silica gel usingethyl acetate in hexane (0-100%) gave the desired compound as a racemicmixture, 1.2 g, 69%. LCMS calculated for C₁₇H₂₄ClNO₅Na (M+Na)⁺:m/z=380.1; found: 380.1. ¹H NMR (500 MHz, DMSO-d₆): δ 7.48 (s, 1H), 6.80(m, 1H), 5.50 (br s, 1H), 5.20 (br s, 1H), 3.83 (s, 3H), 3.32 (m, 1H),3.22 (m, 1H), 2.59 (s, 3H), 2.55 (s, 3H), 1.32 (s, 9H).

Chiral purification on ChiralPak AD-H, 20×250 mm (Chiral Technologies),5 micron particle size, at flow rate of 18 mL/min using 8% ethanol inhexanes gave the Peak 1 (single enantiomer) (retention time=9.86minutes) and Peak 2 (single enantiomer) (retention time=11.47 minutes).

Step 3.N-[2-(3-Acetyl-5-chloro-2-methoxy-6-methylphenyl)-2-hydroxyethyl]-2-chloroacetamide

tert-Butyl[2-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-2-hydroxyethyl]carbamate(170 mg, 0.47 mmol) (Peak 1 from step 2) was treated with 4.0 M hydrogenchloride in 1,4-dioxane (12 mL) for 15 minutes. The solvents wereevaporated, methylene chloride (6 mL) and triethylamine (200 μL, 1.4mmol) were added and the mixture cooled to 0° C. Chloroacetyl chloride(45 μL, 0.56 mmol) was added slowly and was stirred for 10 minutes at 0°C. The solvents were evaporated to dryness. Water was added and themixture was extracted with ethyl acetate. The combined extracts werewashed with brine, dried over sodium sulfate, and concentrated to givethe crude residue as a single enantiomer. LCMS calculated forCl₄H₁₇Cl₂NO₄Na (M+Na)⁺: m/z=356.1; found: 356.1.

Step 4. 6-(3-Acetyl-5-chloro-2-methoxy-6-methylphenyl) morpholin-3-one

To a solution ofN-[2-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-2-hydroxyethyl]-2-chloroacetamide(170 mg, 0.50 mmol) (single enantiomer from step 3) in tetrahydrofuran(4 mL) cooled at 0° C., a mixture of sodium hydride (60% dispersion inmineral oil; 39 mg, 1.0 mmol) was added and stirred for 1 hour. Thereaction was quenched with water and extracted with ethyl acetate. Thecombined extracts were washed with brine, dried over sodium sulfate, andconcentrated to give the crude residue as a single enantiomer, 61 mg,41%. LCMS calculated for C₁₄H₁₇ClNO₄ (M+H)⁺: m/z=298.1; found: 298.1.

Step 5.6-[3-Chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]morpholin-3-one

To a solution of6-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)morpholin-3-one (27 mg,0.090 mmol) (single enantiomer from step 4) in methanol (2 mL) was addedsodium tetrahydroborate (6.8 mg, 0.18 mmol) at 0° C. and stirred for 1hour. Purification by preparative LCMS (pH 10) gave the desired compoundas a racemic mixture of two diastereomers, 20 mg, 76%. LCMS calculatedfor C₁₄H₁₇ClNO₃ (M-OH)⁺: m/z=282.1; found: 282.1.

Step 6.6-[3-Chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]morpholin-3-one

A mixture of thionyl chloride (15 μL, 0.21 mmol) andN,N-dimethylformamide (10.0 μL) was stirred at room temperature for 10minutes. A solution of6-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]morpholin-3-one(19.0 mg, 0.0634 mmol) (racemic mixture of two diastereomers from step5) in methylene chloride (1.0 mL) was added and the mixture was stirredat room temperature overnight. The mixture was diluted with methylenechloride, washed with saturated sodium bicarbonate, water, brine, driedover sodium sulfate, filtered and concentrated to give the desiredcompound as a racemic mixture of two diastereomers, 19 mg, 94%. LCMScalculated for C₁₄H₁₇ClNO₃ (M-C1)⁺: m/z=282.1; found: 282.1.

Step 7.6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}morpholin-3-one

A mix of6-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]morpholin-3-one(19.0 mg, 0.0597 mmol) (racemic mixture of two diastereomers from step6) 3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (11 mg, 0.072 mmol),cesium carbonate (29 mg, 0.090 mmol) and potassium iodide (0.99 mg,0.006 mmol) in N,N-dimethylformamide (0.19 mL) was heated at 140° C. for1 hour. The mixture was diluted with ether, washed with water,concentrated and purified by preparative LCMS (pH 10) using RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 30 mL/min) to give2.5 mg, 10% of Peak 1 (single enantiomer, retention time 10.15 min):LCMS calculated for C₂₀H₂₄ClN₆O₃(M+H)⁺: m/z=431.2; found: 431.1, and 2.7mg, 10% of Peak 2 (single enantiomer, retention time 10.76 min): LCMScalculated for C₂₀H₂₄ClN₆O₃(M+H)⁺: m/z=431.2; found: 431.1.

Example 325.5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-1,3-oxazolidin-2-one

Step 1.5-(3-Acetyl-5-chloro-2-methoxy-6-methylphenyl)-1,3-oxazolidin-2-one

To a solution of tert-butyl[2-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-2-hydroxyethyl]carbamate(140 mg, 0.40 mmol) (Peak 1, single enantiomer from step 2, Example 324)in tetrahydrofuran (2.5 mL), N,N-diisopropylethylamine (0.35 mL, 2.0mmol) and N,N-carbonyldiimidazole (130 mg, 0.80 mmol). The reaction wasrefluxed at 70° C. for 10 minutes. The reaction was evaporated todryness. Purification on silica gel using (0-50%) ethyl acetate inhexane gave the desired compound as a single enantiomer, 78 mg, 69%.LCMS calculated for C₁₃H₁₅ClNO₄ (M+H)⁺: m/z=284.1; found: 284.1.

Step 2.5-[3-Chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]-1,3-oxazolidin-2-one

To a solution of5-(3-acetyl-5-chloro-2-methoxy-6-methylphenyl)-1,3-oxazolidin-2-one (21mg, 0.072 mmol) (single enantiomer from step 1) in methanol (1 mL) wasadded sodium tetrahydroborate (5.5 mg, 0.14 mmol) at 0° C. The mixturewas stirred at 0° C. for 1 hour. It was diluted with methanol andpurified on preparative LCMS using pH 10 buffer to give the desiredcompound as a racemic mixture of two diastereomers, 17 mg, 83%. LCMScalculated for C₁₃H₁₅ClNO₃ (M-OH)⁺: m/z=268.1; found: 268.1.

Step 3.5-[3-Chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]-1,3-oxazolidin-2-one

A mixture of cyanuric chloride (16 mg, 0.084 mmol) andN,N-dimethylformamide (15 μL) was stirred at room temperature for 10minutes. A solution of5-[3-chloro-5-(1-hydroxyethyl)-6-methoxy-2-methylphenyl]-1,3-oxazolidin-2-one(16 mg, 0.056 mmol) (racemic mixture of two diastereomers from step 2)in methylene chloride (0.3 mL) was added and the reaction was stirred atroom temperature overnight. Thionyl chloride (12 μL, 0.17 mmol) wasadded and stirred for 10 min. The mixture was diluted with methylenechloride, washed with saturated sodium bicarbonate, water, brine, driedover sodium sulfate, filtered and concentrated to give the desiredcompound as a racemic mixture of two diastereomers, 17 mg, 100%. LCMScalculated for C₁₃H₁₆Cl₂NO₃ (M+H)⁺: m/z=304.0; found: 304.1.

Step 4.5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-1,3-oxazolidin-2-one

A mixture of5-[3-chloro-5-(1-chloroethyl)-6-methoxy-2-methylphenyl]-1,3-oxazolidin-2-one(17 mg, 0.056 mmol) (racemic mixture of two diastereomers from step 3)3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (10 mg, 0.067 mmol), cesiumcarbonate (27 mg, 0.084 mmol) and potassium iodide (0.93 mg, 0.0056mmol) in N,N-dimethylformamide (0.18 mL) was heated at 140° C. for 1hour. The mixture was diluted with ether, washed with water,concentrated and purified by preparative LCMS (pH 10) to give thedesired compound as a racemic mixture of two diastereomers, 2.2 mg, 9%;LCMS calculated for C₁₉H₂₂ClN₆O₃(M+H)⁺: m/z=417.1; found: 417.1.

Examples 345-348. Diastereoisomers of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}pyrrolidin-2-one

Step 1. 1-(5-Chloro-2-ethoxy-3-iodo-4-methylphenyl)ethanol

A solution of 1-(5-chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone (20.0g, 58.4 mmol; Example 212, step 1) and 1,2-ethanediol (6.5 mL, 120 mmol)in toluene (190 mL) was treated with p-toluenesulfonic acid monohydrate(1.1 g, 5.8 mmol). The flask was fitted with a Dean-Stark trap that wasfilled with sieves, and refluxed for 3 h. The reaction mixture wascooled and added to ice cooled saturated sodium bicarbonate solution(250 mL) and extracted with ethyl acetate. The organic layer was washedwith brine, dried over sodium sulfate, filtered, and concentrated to acrude orange oil. The crude material was purified by flash columnchromatography using ethyl acetate in hexanes (0%-20%) to give thedesired product (22 g, 99%). LCMS for C₁₂H₁₄ClFIO₃ (M+H)⁺: m/z=387.0;Found: 386.9.

Step 2. Ethyl(2E)-3-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]acrylate

A mixture of2-(5-chloro-2-ethoxy-4-fluoro-3-iodophenyl)-2-methyl-1,3-dioxolane (22g, 58 mmol) (from Step 1), ethyl(2E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)acrylate (16 mL, 70mmol), and potassium carbonate (24 g, 170 mmol) in 1,4-dioxane (230 mL)and water (110 mL) was degassed with nitrogen for 10 min. The reactionmixture was treated with [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complex withdichloromethane (1:1) (2.4 g, 2.9 mmol), degassed with nitrogen foranother 10 min, and heated at 80° C. for 2 h. The reaction mixture wasfiltered through Celite and washed with ethyl acetate (300 mL). Thefiltrate was poured into water (400 mL). The aqueous layer was separatedand extracted with additional ethyl acetate (300 mL). The combinedorganic extracts were washed with brine, dried over sodium sulfate,filtered, and concentrated to a crude brown solid. The crude materialwas purified by flash column chromatography using ethyl acetate inhexanes (0%-30%) to give the desired product (20 g, 96%). ¹H NMR (400MHz, CDCl₃) δ 7.74 (d, J=16.5 Hz, 1H), 7.56 (d, J=8.6 Hz, 1H), 6.70 (dd,J=16.5, 0.9 Hz, 1H), 4.26 (q, J=7.1 Hz, 2H), 4.10-3.99 (m, 2H), 3.91 (q,J=7.0 Hz, 2H), 3.87-3.76 (m, 2H), 1.73 (s, 3H), 1.44 (t, J=7.0 Hz, 3H),1.33 (t, J=7.1 Hz, 3H). LCMS for C₁₇H₂₁ClFO₅ (M+H)⁺: m/z=359.1; Found:359.1.

Step 3. Ethyl3-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]-4-nitrobutanoate

A solution ethyl(2E)-3-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]acrylate(10 g, 28 mmol) (from Step 2) in nitromethane (100 mL) was treated with1,8-diazabicyclo[5.4.0]undec-7-ene (4.6 mL, 31 mmol) and stirred at 60°C. for 15 h. The reaction mixture was poured into water (400 mL) andextracted with ethyl acetate (2×300 mL). The combined organic extractswere washed with brine, dried over sodium sulfate, filtered, andconcentrated to a crude orange oil. The crude material was purified byflash column chromatography using ethyl acetate in hexanes (0%-30%) togive the desired product as a mixture of enantiomers (10.4 g, 89%). ¹HNMR (400 MHz, CDCl₃) δ 7.52 (d, J=9.1 Hz, 1H), 4.82 (ddd, J=12.5, 7.6,1.4 Hz, 1H), 4.68 (dd, J=12.5, 7.2 Hz, 1H), 4.54-4.40 (m, 1H), 4.15-3.90(m, 6H), 3.89-3.75 (m, 2H), 2.85 (ddd, J=16.0, 8.6, 1.4 Hz, 1H), 2.73(dd, J=16.1, 6.2 Hz, 1H), 1.70 (s, 3H), 1.47 (t, J=7.0 Hz, 3H), 1.21 (t,J=7.1 Hz, 3H). LCMS for C₁₈H₂₄ClFNO₇ (M+H)⁺: m/z=420.1; Found: 420.1.

Step 4. Enantiomers4-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]pyrrolidin-2-one

A suspension of ethyl3-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]-4-nitrobutanoate(1.0 g, 2.4 mmol) (from Step 3) in ethanol (16 mL) was warmed todissolve the solid. The solution was cooled back to ambient temperature,degassed with nitrogen, and treated with a slurry of 2800 Raney Nickelin water (1.5 mL). The reaction mixture was degassed again with nitrogenand hydrogenated with a balloon of hydrogen for 3 h. The reactionmixture was filtered through Celite and concentrated to give theintermediate amino ester (0.93 g, 100%). The intermediate amino esterwas dissolved in toluene (12 mL) and heated at 110° C. for 12 h. Thereaction mixture was cooled to ambient temperature, at which point asolid precipitated from solution. This mixture was cooled to 0° C.,stirred for 30 min, filtered, washed with cold toluene, and dried togive the desired product as a mixture of enantiomers (0.61 g, 75%). LCMSfor C₁₆H₂₀ClFNO₄ (M+H)⁺: m/z=344.1; Found: 344.1. The mixture ofenantiomers was separated by chiral HPLC to give the individualenantiomers as peak 1 and peak 2 (RT=5.39 min and 7.01 min,respectively; Phenomenex Lux Cellulose C-1, 21.2×250 mm, 5 micronparticle size, eluting with 20% ethanol in hexanes at 18 mL/min).

Step 5. Enantiomers of4-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one

The separated enantiomers from step 4 were each processed individuallyto the final compounds. A solution of4-[3-chloro-6-ethoxy-2-fluoro-5-(2-methyl-1,3-dioxolan-2-yl)phenyl]pyrrolidin-2-one(1.7 g, 5.0 mmol) (from Step 4) in methanol (17 mL) was treated with 6.0M hydrogen chloride in water (11 mL, 69 mmol) dropwise and stirred 20°C. for 30 min. The reaction mixture was added dropwise to ice cooledsaturated sodium bicarbonate solution (75 ml) and extracted with ethylacetate (2×100 ml). The combined organic extracts were washed withbrine, dried over sodium sulfate, filtered, and concentrated to give thedesired products [from peak 1 (1.5 g, 99%); from peak 2 (1.5 g, 99%)]that were used without further purification. From peak 1: ¹H NMR (400MHz, DMSO-d₆) δ 7.84 (s, 1H), 7.70 (d, J=8.6 Hz, 1H), 4.16-3.99 (m, 1H),3.83 (q, J=7.0 Hz, 2H), 3.65-3.54 (m, 1H), 3.30-3.23 (m, 1H), 2.55 (s,3H), 2.33 (dd, J=16.8, 8.4 Hz, 1H), 1.30 (t, J=7.0 Hz, 3H). LCMS forC₁₄H₁₆ClFNO₃ (M+H)⁺: m/z=300.1; Found: 300.0. From peak 2: ¹H NMR (400MHz, DMSO-d₆) δ 7.84 (s, 1H), 7.70 (d, J=8.6 Hz, 1H), 4.13-4.00 (m, 1H),3.87-3.77 (m, 2H), 3.65-3.55 (m, 1H), 3.31-3.23 (m, 1H), 2.55 (s, 3H),2.32 (ddd, J=16.9, 8.4, 1.6 Hz, 1H), 1.30 (t, J=7.0 Hz, 3H). LCMS forC₁₄H₁₆ClFNO₃ (M+H)⁺: m/z=300.1; Found: 300.1.

Step 6. Diastereoisomers of4-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]pyrrolidin-2-one

The enantiomers from step 5 were each processed individually to thefinal products. A solution of4-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one (0.402 g,1.34 mmol) (from Step 5) in anhydrous methanol (6.7 mL) under anatmosphere of nitrogen at 0° C. was treated with sodium tetrahydroborate(0.10 g, 2.7 mmol) and stirred at 0° C. for 30 min. The reaction mixturewas quenched with water at 0° C. and poured into water (50 mL)/ethylacetate (100 mL) while stirring. The mixture was warmed to ambienttemperature and the aqueous layer was separated and extracted withadditional ethyl acetate (50 mL). The combined organic extracts werewashed with brine, dried over sodium sulfate, filtered, and concentratedto give white foams. The crude material were purified by flash columnchromatography using acetonitrile (containing 7% methanol) indichloromethane (0%-100%) to give the desired products as mixtures ofdiastereoisomers [from peak 1 (0.40 g, 99%); from peak 2 (0.40 g, 99%)].From peak 1: LCMS for C₁₄H₁₈ClFNO₃ (M+H)⁺: m/z=302.1; Found: 302.0. Frompeak 2: LCMS for C₁₄H₁₈ClFNO₃ (M+H)⁺: m/z=302.1; Found: 302.1.

Step 7. Diastereoisomers of4-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-fluorophenyl]pyrrolidin-2-one

The mixture of diastereoisomers from step 6 were each processedindividually to the final products. A solution of4-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]pyrrolidin-2-one(0.41 g, 1.4 mmol) (from Step 6) in methylene chloride (12 mL) wastreated with N,N-dimethylformamide (0.011 mL, 0.14 mmol) followed bythionyl chloride (0.21 mL, 2.9 mmol) dropwise and stirred at 20° C. for30 min. The reaction mixture was added dropwise to ice cooled saturatedsodium bicarbonate solution and extracted with dichloromethane. Theorganic layer was separated and washed with brine, dried over sodiumsulfate, filtered, and concentrated to give the desired products [frompeak 1 (0.38 g, 87%); from peak 2 (0.39 g, 89%)] along with 17-18% ofthe styrene that formed from chloride elimination. These mixtures wereused without further purification. From peak 1: LCMS for C₁₄H₁₇Cl₂FNO₂(M+H)⁺: m/z=320.1; Found: 320.0. From peak 2: LCMS for C₁₄H₁₇Cl₂FNO₂(M+H)⁺: m/z=320.1; Found: 320.0.

Step 8. Diastereoisomers of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}pyrrolidin-2-one

The mixture of diastereoisomers from step 7 were each processedindividually to the final products. A mixture of4-[3-chloro-5-(1-chloroethyl)-6-ethoxy-2-fluorophenyl]pyrrolidin-2-one(0.36 g, 1.1 mmol) (from Step 7),3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.19 g, 1.3 mmol), cesiumcarbonate (0.54 g, 1.7 mmol) and potassium iodide (18 mg, 0.11 mmol) inN,N-dimethylformamide (7.4 mL) was heated at 100° C. for 4.5 h. Thereaction mixture was poured into water (30 ml) and extracted with ethylacetate (3×50 mL) to give a mixture of diastereoisomer((S)-4-(3-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one;(R)-4-(3-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one;(S)-4-(3-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one;and(R)-4-(3-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one).The mixture of diastereoisomers were purified by preparative LCMS(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 60 mL/min) to givethe desired products [from peak 1 were isolated peak A (compound 345)(0.13 g, 54%) and peak B (compound 346) (0.11 g, 46%); from peak 2 wereisolated peak A (compound 347) (0.15 g, 63%) and peak B (compound 348)(0.14 g, 55%)]. Compound 346: ¹H NMR (300 MHz, DMSO-d₆) δ 8.12 (s, 1H),7.82 (s, 1H), 7.52 (d, J=8.5 Hz, 1H), 7.30 (br s, 1H), 6.23 (q, J=7.0Hz, 1H), 4.05-3.90 (m, 1H), 3.88-3.78 (m, 2H), 3.63-3.53 (m, 1H),3.29-3.20 (m, 1H), 2.54 (s, 3H), 2.38-2.21 (m, 1H), 1.70 (d, J=7.1 Hz,3H), 1.39 (t, J⁼6.9 Hz, 3H). LCMS for C₂₀H₂₃ClFN₆O₂(M+H)⁺: m/z=433.2;Found: 433.1. Compound 347: ¹H NMR (500 MHz, DMSO-d₆) δ 8.12 (s, 1H),7.77 (s, 1H), 7.53 (d, J=8.5 Hz, 1H), 7.26 (br s, 2H), 6.24 (q, J=7.0Hz, 1H), 4.04-3.94 (m, 1H), 3.93-3.85 (m, 1H), 3.84-3.77 (m, 1H),3.61-3.53 (m, 1H), 3.27-3.22 (m, 1H), 2.54 (s, 3H), 2.30 (dd, J=18.1,8.6 Hz, 1H), 1.71 (d, J=7.1 Hz, 3H), 1.40 (t, J=6.9 Hz, 3H). LCMS forC₂₀H₂₃ClFN₆O₂(M+H)⁺: m/z=433.2; Found: 433.1.

Examples 349-352. Diastereoisomers of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile

Step 1. Enantiomers of4-acetyl-6-chloro-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile

A racemic mixture of4-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one (0.20 g,0.67 mmol) (from Example 345, Step 5) and sodium cyanide (0.057 g, 1.2mmol) in dimethyl sulfoxide (1.5 mL) was stirred at 80° C. for 3 h. Thereaction mixture was poured into water (35 mL) and extracted with ethylacetate (2×50 mL). The combined organic extracts were washed with brine,dried over sodium sulfate, filtered, and concentrated to give a cruderesidue. The crude material was purified by flash column chromatographyusing ether (containing 10% methanol) in hexanes (0%-100%) to give thedesired product (0.15 g, 71%) as a mixture of enantiomers. LCMS forC₁₅H₁₆ClN₂O₃(M+H)⁺: m/z=307.1; Found: 307.0. The mixture of enantiomerswas separated by chiral HPLC to give the individual enantiomers as peak1 and peak 2 (RT=5.00 min and 10.4 min; Phenomenex Lux Cellulose C-2,21.2×250 mm, 5 micron particle size, eluting with 60% ethanol in hexanesat 18 mL/min).

Step 2. Diastereoisomers of6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(5-oxopyrrolidin-3-yl)benzonitrile

The enantiomers from step 1 were each processed individually to thefinal products. A solution of4-acetyl-6-chloro-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile (frompeak 1: 0.83 g, 2.7 mmol; from peak 2: 0.86 g, 2.8 mmol) in anhydrousmethanol (14 mL) under an atmosphere of nitrogen at 0° C. was treatedwith sodium tetrahydroborate (0.20 g, 5.4 mmol) and stirred at 0° C. for30 min. The reaction mixture was quenched with water at 0° C. and pouredinto water (50 mL)/ethyl acetate (100 mL) while stirring. The mixturewas warmed to ambient temperature and the aqueous layer was separatedand extracted with additional ethyl acetate (50 mL). The combinedorganic extracts were washed with brine, dried over sodium sulfate,filtered, and concentrated to give the desired products as mixtures ofdiastereoisomers [from peak 1 (0.83 g, 99%); from peak 2 (0.87 g, 99%)].From peak 1: LCMS for C₁₅H₁₈ClN₂O₃(M+H)⁺: m/z=309.1; Found: 309.1. Frompeak 2: LCMS for C₁₅H₁₈ClN₂O₃(M+H)⁺: m/z=309.1; Found: 309.1.

Step 3. Diastereoisomers of6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile

The mixture of diastereoisomers from step 2 were each processedindividually to the final products. A solution of6-chloro-3-ethoxy-4-(1-hydroxyethyl)-2-(5-oxopyrrolidin-3-yl)benzonitrile(from peak 1: 0.83 g, 2.7 mmol; from peak 2: 0.87 g, 2.8 mmol) inmethylene chloride (23 mL) was treated with N,N-dimethylformamide (0.021mL, 0.27 mmol) followed by thionyl chloride (0.490 mL, 6.72 mmol)dropwise and stirred at 20° C. for 2 h. The reaction mixture was addeddropwise to ice cooled saturated sodium bicarbonate solution andextracted with dichloromethane. The organic layer was separated andwashed with brine, dried over sodium sulfate, filtered, and concentratedto give the desired products as mixtures of diastereoisomers [from peak1 (0.85 g, 97%); from peak 2 (0.90 g, 98%)]. These mixtures were usedwithout further purification. From peak 1: LCMS for C₁₅H₁₇Cl₂N₂O₂(M+H)⁺: m/z=327.1; Found: 327.1. From peak 2: LCMS for C₁₅H₁₇Cl₂N₂O₂(M+H)⁺: m/z=327.1; Found: 327.1.

Step 4. Diastereoisomers of4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile

The mixture of diastereoisomers from step 3 were each processedindividually. A mixture of6-chloro-4-(1-chloroethyl)-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile(from peak 1: 0.85 g, 2.6 mmol; from peak 2: 0.89 g, 2.7 mmol),3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (0.46 g, 3.1 mmol), cesiumcarbonate (1.3 g, 3.9 mmol) and potassium iodide (43 mg, 0.26 mmol) inN,N-dimethylformamide (17 mL, 220 mmol) was heated at 90° C. for 3 h.The reaction mixture was poured into water (100 mL)/ethyl acetate (100mL) and filtered through Celite to remove black solids. The aqueouslayer was separated and extracted with ethyl acetate (2×100 mL). Thecombined organic extracts were washed with brine, dried over sodiumsulfate, filtered, and concentrated to give white foams. The crudematerial were purified by flash column chromatography using methanol indichloromethane (0%-20%) to give the desired products as mixtures ofdiastereoisomers [from peak 1 (0.49 g, 43%); from peak 2 (0.53 g, 44%)].Analytical chiral HPLC analysis of the diastereoisomers from peak 1revealed a mixture of four peaks instead of the desired two due toepimerization. Analysis of the diastereoisomers from peak 2 alsorevealed four peaks. Both sets of mixtures were combined and purifiedvia chiral HPLC to give four individual peaks (RT=6.41 min, 8.13 min,9.93 min, 14.4 min; Phenomenex Lux Cellulose C-2, 21.2×250 mm, 5 micronparticle size, eluting with 60% ethanol in hexanes at 18 mL/min). Thecompounds of peak 1 (compound 351), peak 2 (compound 349), peak 3(compound 352), and peak 4 (compound 350) were then tested in the assaysof Example A3 and B2. Compound 349: ¹H NMR (500 MHz, DMSO-d₆) δ 8.12 (s,1H), 7.88 (s, 1H), 7.58 (s, 1H), 7.30 (br s, 2H), 6.26 (q, J=7.0 Hz,1H), 4.32-4.20 (m, 1H), 4.00-3.91 (m, 1H), 3.90-3.81 (m, 1H), 3.65-3.59(m, 1H), 3.49-3.42 (m, 1H), 2.55 (s, 3H), 1.74 (d, J=7.0 Hz, 3H), 1.43(t, J=6.9 Hz, 3H). LCMS for C₂₁H₂₃ClN₇O₂(M+H)⁺: m/z=440.2; Found: 440.2.Compound 352: ¹H NMR (500 MHz, DMSO-d₆) δ 8.12 (s, 1H), 7.88 (s, 1H),7.56 (s, 1H), 7.30 (br s, 2H), 6.26 (q, J=7.0 Hz, 1H), 4.32-4.19 (m,1H), 3.97-3.82 (m, 2H), 3.67-3.59 (m, 1H), 3.49-3.40 (m, 1H), 2.59-2.52(m, 3H), 1.73 (d, J=7.0 Hz, 3H), 1.42 (t, J=6.9 Hz, 3H). LCMS forC₂₁H₂₃ClN₇O₂(M+H)⁺: m/z=440.2; Found: 440.2.

Examples 353 and 354. Diastereomers of4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one

Step 1: 1-(5-Chloro-2-ethoxy-4-fluoro-3-vinylphenyl)ethanone

A mixture of 1-(5-chloro-2-ethoxy-4-fluoro-3-iodophenyl)ethanone (13.3g, 38.8 mmol) (from Example 139, Step 1),4,4,5,5-tetramethyl-2-vinyl-1,3,2-dioxaborolane (7.9 mL, 46 mmol),[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II), complexwith dichloromethane (1:1) (1.0 g, 1.0 mmol) and potassium carbonate (16g, 120 mmol) in 1,4-dioxane (200 mL) and water (100 mL) was heated at80° C. for 2 hours. The mixture was cooled to rt and extracted withethyl acetate. The extracts were washed with brine, dried over sodiumsulfate, filtered and concentrated. Purification on silica gel usingethyl acetate in hexanes (0-30%) gave the desired compound, 7.0 g, 74%.LCMS calculated for C₁₂H₁₃ClFO₂ (M+H)⁺: m/z=243.0; found: 243.1.

Step 2:1-[5-Chloro-3-(1,2-dihydroxyethyl)-2-ethoxy-4-fluorophenyl]ethanone

AD-mix-alpha (5.8 g, 7.3 mmol) (Aldrich #392758) was stirred intert-butyl alcohol (21 mL) with water (21 mL) for 15 minutes.1-(5-chloro-2-ethoxy-4-fluoro-3-vinylphenyl)ethanone (1.0 g, 4.1 mmol)(from Step 1) was added and the suspension was stirred for 16 hours.Sodium sulfite (6.2 g, 49 mmol) was added and the suspension was stirredfor 15 minutes. The reaction mixture was extracted with ethyl acetate.The extracts were washed with brine and dried over sodium sulfate,filtered and evaporated. Purification on silica gel using ethyl acetatein hexanes (0-80%) gave the desired compound as a racemic mixture, 900mg, 80%. Chiral purification on Phenomenex Lux Cellulose C-2, 21.2×250mm (Chiral Technologies), 5 micron particle size, at flow rate of 18mL/min using 20% ethanol in hexanes gave peak 1 (single enantiomer)(retention time=7.88 minutes) and peak 2 (single enantiomer) (retentiontime=11 minutes); the desired enantiomer was peak 2. LCMS calculated forC₁₂H₁₃ClFO₃ (M-OH)⁺: m/z=259.1; found: 259.1.

Step 3:1-[3-(2-{[tert-Butyl(dimethyl)silyl]oxy}-1-hydroxyethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone

1-[5-Chloro-3-(1,2-dihydroxyethyl)-2-ethoxy-4-fluorophenyl]ethanone (700mg, 2 mmol) (from Step 2, peak 2) was stirred in 1,2-dichloroethane (6mL) with N,N-diisopropylethylamine (4.0 mL, 23 mmol) and a 1.0 Msolution of tert-butyldimethylsilyl chloride in 1,2-dichloroethane (7.6mL) was added. The mixture was heated to 80° C. for 3 hours and cooledto rt. Evaporation and purification on silica gel using ethyl acetate inhexanes (0-50%) gave the desired compound 800 mg, 80%. LCMS calculatedfor C₁₈H₂₈ClFO₄SiNa (M+Na)⁺: m/z=413.1; found: 413.1.

Step 4:1-(3-Acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-{[tert-butyl(dimethyl)silyl]oxy}ethylmethanesulfonate

1-[3-(2-{[tert-Butyl(dimethyl)silyl]oxy}-1-hydroxyethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone(700 mg, 2.0 mmol) (from Step 3) was stirred in 1,2-dichloroethane (15mL) with triethylamine (2.0 mL, 14 mmol) and methanesulfonic anhydride(670 mg, 3.8 mmol) at rt for 1.5 hours. The mixture was poured intobrine and extracted with dichloromethane. The extracts were dried oversodium sulfate, filtered and evaporated to give the desired compound 830mg, 100%. LCMS calculated for C₁₈H₂₇ClFO₃Si (M-OMs)⁺: m/z=373.1; found:373.1.

Step 5:1-[3-(1-Azido-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone

1-(3-Acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-{[tert-butyl(dimethyl)silyl]oxy}ethylmethanesulfonate (0.83 g, 1.77 mmol) (from Step 4) was stirred indimethyl sulfoxide (10 mL) and sodium azide (0.12 g, 1.8 mmol) wasadded. The mixture was heated to 50° C. for 1 hour and cooled to rt. Themixture was poured into brine and extracted with ethyl acetate. Theextracts were dried over sodium sulfate, filtered and evaporated to givethe desired compound 736 mg, 100%. LCMS calculated for C₁₈H₂₇ClFN₃O₃SiNa(M+Na)⁺: m/z=438.1; found: 438.1.

Step 6:1-[3-(1-Amino-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone

1-[3-(1-Azido-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone(750 mg, 1.8 mmol) (from Step 5) was stirred in tetrahydrofuran (10 mL)with water (0.33 mL) and triphenylphosphine was added. The mixture washeated to 60° C. for 2 hours and cooled to rt. Brine was added and themixture was extracted with ethyl acetate. The extracts were dried oversodium sulfate, filtered and evaporated to give the desired compound 700mg, 100%. LCMS calculated for C₁₈H₃₀ClFNO₃Si (M+H)⁺: m/z=390.2; found:390.2.

Step 7: tert-Butyl(1-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)carbamate

1-[3-(1-Amino-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)-5-chloro-2-ethoxy-4-fluorophenyl]ethanone(700 mg, 2.0 mmol) (from Step 6) was stirred in tetrahydrofuran (30 mL)with di-tert-butyldicarbonate (780 mg, 3.6 mmol) andN,N-diisopropylethylamine (0.94 mL, 5.4 mmol) was added. The mixture wasstirred at rt for 30 minutes. Brine was added and the mixture wasextracted with ethyl acetate. The extracts were dried over sodiumsulfate, filtered and evaporated. Purification on silica gel using ethylacetate in hexanes (0-30%) gave the desired compound 550 mg, 60%. LCMScalculated for C₂₃H₃₇ClFNO₅SiNa (M+Na)⁺: m/z=512.2; found: 512.2.

Step 8: tert-Butyl[1-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-hydroxyethyl]carbamate

Tert-Butyl(1-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-{[tert-butyl(dimethyl)silyl]oxy}ethyl)carbamate(500 mg, 1.0 mmol) (from Step 7) was stirred in tetrahydrofuran (10 mL)and a 1.0 M solution of tetra-n-butylammonium fluoride intetrahydrofuran (1.5 mL) was added. The mixture was stirred at rt for 30minutes and evaporated. Purification on silica gel using ethyl acetatein hexanes (0-50%) gave the desired compound 238 mg, 60%. LCMScalculated for C₁₇H₂₃ClFNO₅Na (M+Na)⁺: m/z=398.1; found: 398.1.

Step 9:4-(3-Acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-1,3-oxazolidin-2-one

tert-Butyl[1-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-hydroxyethyl]carbamate(234 mg, 0.62 mmol) (from Step 8) was dissolved in 1,2-dichloroethane(12 mL) and a solution of 2.0 M phosgene in toluene (0.93 mL) was added.The mixture was heated to 80° C. for 1.5 hours. Evaporation andpurification on silica gel using ethyl acetate in hexanes (0-85%) gavethe desired compound, 175 mg, 93%. LCMS calculated for C₁₃H₁₄ClFNO₄(M+H)⁺: m/z=302.1; found: 302.1.

Step 10:4-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]-1,3-oxazolidin-2-one

4-(3-Acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-1,3-oxazolidin-2-one (175mg, 0.58 mmol) was stirred in methanol (10 mL) at 0° C. and sodiumtetrahydroborate (33 mg, 0.87 mmol) was added. The mixture was stirredat rt for 1 hour and evaporated. Water was added and the mixture wasextracted with ethyl acetate. The extracts were washed with brine, driedover sodium sulfate, filtered and evaporated to give an approximate 1:1mixture of diastereomers, 175 mg, 99%. LCMS calculated forC₁₃H₁₅ClFNO₄Na (M+Na)⁺: m/z=326.1; found: 326.1.

Step 11:4-[3-chloro-5-(chloroethyl)-6-ethoxy-2-fluorophenyl]-1,3-oxazolidin-2-one

4-[3-chloro-6-ethoxy-2-fluoro-5-(1-hydroxyethyl)phenyl]-1,3-oxazolidin-2-one(150 mg, 0.49 mmol) (from Step 10) was stirred in dichloromethane (4 mL)with N,N-dimethylformamide (96 μL) and thionyl chloride (110 μL, 1.5mmol) was added. The mixture was evaporated. Water was added and themixture was extracted with ethyl acetate. The extracts were washed withbrine, dried over sodium sulfate, filtered and evaporated to give thedesired compound, 159 mg, 100%.

Step 12:4-{3-[1-(4-amino-3-methyl-H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one

4-[3-chloro-5-(chloroethyl)-6-ethoxy-2-fluorophenyl]-1,3-oxazolidin-2-one(160 mg, 0.50 mmol) (from Step 11) was stirred in N,N-dimethylformamide(21 mL) with cesium carbonate (324 mg, 0.99 mmol) and3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine (89 mg, 0.60 mmol) wasadded. The mixture was heated to 80° C. for 1.5 hours and cooled to rt.The mixture was diluted with water and extracted with ethyl acetate. Theextracts were washed with brine, dried over sodium sulfate, filtered andevaporated. Purification by preparative LCMS (pH 10) using RP-HPLC(XBridge C18 column, eluting with a gradient of acetonitrile/watercontaining 0.1% ammonium hydroxide, at flow rate of 30 mL/min) separatedthe two diastereomers (peak 1 [compound 353] Rt=4.9 min. and peak 2[compound 354] Rt=5.6 min.); providing compound 354 as the desiredsingle enantiomer, 28 mg, 13%. peak 2: LCMS calculated for C₁₉H₂₁ClFN₆O₃(M+H)⁺: m/z=435.1; found: 435.1. ¹H NMR (300 MHz, CD₃OD): δ 8.15 (s,1H), 7.62 (m, 1H), 6.31 (m, 1H), 5.39 (m, 1H), 4.79 (m, 1H), 4.40 (m,1H), 3.95 (m, 1H), 3.80 (m, 1H), 2.60 (s, 3H), 1.80 (m, 3H), 1.40 (m,3H).

Examples 355-358. Diastereomers of5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one

Step 1: tert-Butyl[2-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-hydroxyethyl]carbamate

0.2 M Osmium tetraoxide in water (10 mL) was added to a solution oftert-butyl [(4-chlorobenzoyl)oxy]carbamate (Lawrence Harris, J Org.Chem, 2011, 76, 358-372). (19 g, 70 mmol) in acetonitrile (210 mL) andstirred for 10 minutes.1-(5-chloro-2-ethoxy-4-fluoro-3-vinylphenyl)ethanone (11.2 g, 46 mmol)(from Example 353, Step 1) as a solution in acetonitrile (210 mL) wasadded to the carbamate solution followed by the addition of water (50mL) and the reaction was stirred for 3 hours at room temperature. Thereaction was quenched with saturated 10 M dipotassium disulfite in water(240 mL) and stirred for 5 minutes. Water was added and the reactionmixture was extracted with ethyl acetate. The extracts were washed withsaturated sodium bicarbonate solution, brine and dried over sodiumsulfate, filtered and evaporated. Purification on silica gel using ethylacetate in hexanes (0-100%) gave the desired compound as a racemicmixture, 16.6 g, 95%. LCMS calculated for C₁₇H₂₃ClFNO₅Na (M+Na)⁺:m/z=398.1; found: 398.0.

Step 2:5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one

The desired single enantiomer (peak 3) was prepared using the sameprocedure as Example 353 (steps 8-12), except that the intermediate fromstep 1 in this example was racemic and thus the final separation of thefour diastereomers occurred in step 12. Chiral purification onPhenomenex Lux Cellulose C-4, 21×250 mm (Chiral Technologies), 5 micronparticle size, at flow rate of 18 mL/min using 30% ethanol in hexanesgave the peak 1: compound 355 (single enantiomer) (retention time=12.7minutes), peak 2: compound 356 (single enantiomer) (retention time=14.2minutes), peak 3: compound 357 (single enantiomer) (retention time=20.3minutes), and peak 4: compound 358 (single enantiomer) (retentiontime=28.9 minutes); the most active enantiomer was peak 3. LCMScalculated for C₁₉H₂₁ClFN₆O₃(M+H)⁺: m/z=435.1; found: 435.1. ¹H NMR (500MHz, DMSO-d₆): δ 8.15 (s, 1H), 7.81 (s, 1H), 7.71 (d, 1H), 7.26 (bs,1H), 6.23 (m, 1H), 5.84 (t, 1H), 3.92 (m, 1H), 3.83 (m, 1H), 2.52 (s,3H), 1.75 (d, 3H), 1.40 (m, 3H).

Examples 361-363. Diastereomers of4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-(2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile

Based on the stereochemistry of Example 269, the stereochemistry of eachdiastereomer is believed to be4-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-((S)-2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile(Example 361),4-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-((R)-2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile(Example 362), and4-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-((R)-2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile(Example 363) (structures shown below)

Synthesis of Example 361

To(R)-4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-2-(azetidin-3-yl)-6-chloro-3-methoxybenzonitrile(6.00 g, 14.3 mmol) was added methanol (72 mL). To the resultingsuspension was added (S)-(−)-methyloxirane (2.01 mL, 28.6 mmol) at roomtemperature and the mixture was stirred at room temperature for 19 h.Additional (S)-(−)-methyloxirane (0.50 mL, 7.2 mmol) was added and thestirring was continued for an additional hour. To the reaction mixturewas added water (280 mL) and the cloudy solution was stirred. Themixture was extracted with methylene chloride (300 mL×4). The organiclayer was combined and washed with brine (50 mL) and concentrated. Thecrude product was purified by silica column chromatography eluted withMeOH (contained about 0.5% ammonium hydroxide) in methylene chloride.The fractions contained product were collected and evaporated todryness. This residue was further purified by preparative HPLC to givethe title compound. A sample of the title compound was analyzed by NMRspectroscopy and mass spectrometry and gave the following data. ¹H NMR(500 MHz, DMSO) δ 8.11 (s, 1H), 7.47 (s, 1H), 7.30 (br s, 2H), 6.24 (q,J=7.0 Hz, 1H), 4.32 (br s, 1H), 4.07 (m, 1H), 3.94 (m, 2H), 3.65 (s,3H), 3.59 (m, 1H), 3.08 (m, 2H), 2.56 (s, 3H), 2.38-2.19 (m, 2H), 1.73(d, J=7.1 Hz, 3H), 1.00 (d, J=6.2 Hz, 3H) ppm. LCMS forC₂₂H₂₇ClN₇O₂(M+H)⁺: m/z=456.2; found: 456.2.

Synthesis of Example 362

To(S)-4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-2-(azetidin-3-yl)-6-chloro-3-methoxybenzonitrile(293.0 mg, 0.73 mmol) was added methanol (3.7 mL). To the resultingsuspension was added (R)-(+)-methyloxirane 103 μL, 1.46 mmol) at roomtemperature and the mixture was stirred at room temperature for 19 h.Additional (R)-(+)-methyloxirane (51.3 μL, 0.73 mmol) was added and thestirring was continued for additional 2.5 hours. To the reaction mixturewas added water (14 mL) and the cloudy solution was stirred. The mixturewas extracted with methylene chloride (4×16 mL). The organic layer wascombined and washed with brine (50 mL) and concentrated. The crudeproduct was purified by silica column chromatography, eluted with MeOH(contained about 0.5% ammonium hydroxide) in methylene chloride. Thefractions contained product were collected and evaporated to dryness.This residue was further purified by preparative HPLC to give the titlecompound. A sample of the title compound was analyzed by NMRspectroscopy and mass spectrometry and gave the following data. ¹H NMR(500 MHz, DMSO) δ 8.11 (s, 1H), 7.47 (s, 1H), 7.30 (br s, 2H), 6.24 (q,J=7.0 Hz, 1H), 4.37 (br s, 1H), 4.09 (m, 2H), 3.93 (m, 2H), 3.65 (s,3H), 3.59 (m, 1H), 3.12 (m, 2H), 2.56 (s, 3H), 2.39-2.26 (m, 2H), 1.73(d, J=7.1 Hz, 3H), 1.00 (d, J=6.2 Hz, 3H) ppm. LCMS forC₂₂H₂₇ClN₇O₂(M+H)⁺: m/z=456.2; found: 456.2.

Synthesis of Example 363

To(R)-4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-2-(azetidin-3-yl)-6-chloro-3-methoxybenzonitrile(6.0 g, 14.3 mmol) was added methanol (72 mL). To the resultingsuspension was added (R)-(+)-methyloxirane (2.01 mL, 28.6 mmol) at roomtemperature and the mixture was stirred at room temperature for 18 h. Tothe reaction mixture was added water (280 mL) and the cloudy solutionwas stirred. The mixture was extracted with methylene chloride (300mL×4). The organic layer was combined and washed with brine (50 mL) andconcentrated. The crude product was purified by silica columnchromatography, eluted with MeOH (contained about 0.5% ammoniumhydroxide) in methylene chloride. The fractions contained product werecollected and evaporated to dryness. This residue was further purifiedby preparative HPLC to give the title compound. A sample of the titlecompound was analyzed by NMR spectroscopy and mass spectrometry and gavethe following data. ¹H NMR (500 MHz, DMSO) δ 8.11 (s, 1H), 7.46 (s, 1H),7.29 (br s, 2H), 6.24 (q, J=7.0 Hz, 1H), 4.31 (d, J=4.2 Hz, 1H),4.11-4.00 (m, 1H), 3.98-3.90 (m, 1H), 3.65 (s, 3H), 3.61-3.53 (m, 2H),3.07 (m, 2H), 2.56 (s, 3H), 2.28 (d, J=5.9 Hz, 2H), 1.73 (d, J=7.1 Hz,3H), 1.00 (d, J=6.2 Hz, 3H) ppm.

Three HPLC methods were developed to separate the stereoisomers from thecompound of Example 269. Method A was developed to separate thediastereomer Example 361 from Example 269. The retention times ofExample 361 from Example 269 are 15.7 min and 11.5 min respectively.Chromatographic conditions are described in Table B1.

TABLE B1 Column Phenomenex Cellulose 3 (250 mm, 4.6 mm, 5 micron) MobilePhase 89.9% hexane/10% ethanol/0.1% diethylamine (pre-mixed) Flow Rate 1 mL/min Run Time  30 min Detection Wavelength 247 nm Quantitation Peakarea ratio

Method B was developed to separate the diastereomer Example 362 fromExample 269. The retention times of Example 362 from Example 269 are26.4 min and 21.7 min respectively. Chromatographic conditions aredescribed in Table B2.

TABLE B2 Column Phenomenex Cellulose 4 (250 mm, 4.6 mm, 5 micron) MobilePhase 84.9% hexane/15% ethanol/0.1% diethylamine (pre-mixed) Flow Rate 1 mL/min Run Time  40 min Detection Wavelength 247 nm Quantitation Peakarea ratio

Method C was developed to separate the three stereoisomers Example 361,Example 362 and Example 363 from Example 269. The stereoisomers Example361, Example 362 and Example 363 elute at retention time 12.9 min as abroad band while Example 269 elutes at retention time 14.3 min. Anestimation of the level of the enantiomer, Example 363 can be made by acombination of data from Methods A, B, and C. Chromatographic conditionsare described in Table B3.

TABLE B3 Column Phenomenex Cellulose 1 (250 mm, 4.6 mm, 5 micron) MobilePhase 88% hexanes, 12% ethanol (conatins 0.1% diethylamine) Flow Rate  1mL/min Run Time  25 min Detection Wavelength 247 nm Quantitation Peakarea ratio

Example A1: PI3K Enzyme Assay

PI3-Kinase luminescent assay kit including lipid kinase substrate,D-myo-phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2)D(+)-sn-1,2-di-O-octanoylglyceryl, 3-O-phospho linked (PIP2),biotinylated I(1,3,4,5)P4, PI(3,4,5)P3 Detector Protein is purchasedfrom Echelon Biosciences (Salt Lake City, Utah). AlphaScreen™ GSTDetection Kit including donor and acceptor beads was purchased fromPerkinElmer Life Sciences (Waltham, Mass.). PI3Kδ (p110δ/p85α) ispurchased from Millipore (Bedford, Mass.). ATP, MgCl₂, DTT, EDTA, HEPESand CHAPS are purchased from Sigma-Aldrich (St. Louis, Mo.).

AlphaScreen™ Assay for PI3Kδ

The kinase reaction are conducted in 384-well REMP plate from ThermoFisher Scientific in a final volume of 40 μL. Inhibitors are firstdiluted serially in DMSO and added to the plate wells before theaddition of other reaction components. The final concentration of DMSOin the assay is 2%. The PI3K assays are carried out at room temperaturein 50 mM HEPES, pH 7.4, 5 mM MgCl₂, 50 mM NaCl, 5 mM DTT and CHAPS0.04%. Reactions are initiated by the addition of ATP, the finalreaction mixture consisted of 20 μM PIP2, 20 μM ATP, 1.2 nM PI3Kδ areincubated for 20 minutes. 10 μL of reaction mixture are then transferredto 5 μL 50 nM biotinylated I(1,3,4,5)P4 in quench buffer: 50 mM HEPES pH7.4, 150 mM NaCl, 10 mM EDTA, 5 mM DTT, 0.1% Tween-20, followed with theaddition of 10 μL AlphaScreen™ donor and acceptor beads suspended inquench buffer containing 25 nM PI(3,4,5)P3 detector protein. The finalconcentration of both donor and acceptor beads is 20 mg/ml. After platesealing, the plate are incubated in a dark location at room temperaturefor 2 hours. The activity of the product is determined on Fusion-alphamicroplate reader (Perkin-Elmer). IC₅₀ determination is performed byfitting the curve of percent control activity versus the log of theinhibitor concentration using the GraphPad Prism 3.0 software.

Example A2: PI3K Enzyme Assay

Materials:

Lipid kinase substrate, phosphoinositol-4,5-bisphosphate (PIP2), arepurchased from Echelon Biosciences (Salt Lake City, Utah). PI3K isoformsα, β, δ and γ are purchased from Millipore (Bedford, Mass.). ATP, MgCl₂,DTT, EDTA, MOPS and CHAPS are purchased from Sigma-Aldrich (St. Louis,Mo.).

The kinase reaction are conducted in clear-bottom 96-well plate fromThermo Fisher Scientific in a final volume of 24 μL. Inhibitors arefirst diluted serially in DMSO and added to the plate wells before theaddition of other reaction components. The final concentration of DMSOin the assay is 0.5%. The PI3K assays are carried out at roomtemperature in 20 mM MOPS, pH 6.7, 10 mM MgCl₂, 5 mM DTT and CHAPS0.03%. The reaction mixture is prepared containing 50 μM PIP2, kinaseand varying concentration of inhibitors. Reactions are initiated by theaddition of ATP containing 2.2 μCi [γ-³³P]ATP to a final concentrationof 1000 μM. The final concentration of PI3K isoforms α, β, δ and γ inthe assay were 1.3, 9.4, 2.9 and 10.8 nM, respectively. Reactions areincubated for 180 minutes and terminated by the addition of 100 μL of 1M potassium phosphate pH 8.0, 30 mM EDTA quench buffer. A 100 μL aliquotof the reaction solution are then transferred to 96-well MilliporeMultiScreen IP 0.45 gm PVDF filter plate (The filter plate is prewettedwith 200 μL 100% ethanol, distilled water, and 1 M potassium phosphatepH 8.0, respectively). The filter plate is aspirated on a MilliporeManifold under vacuum and washed with 18×200 μL wash buffer containing 1M potassium phosphate pH 8.0 and 1 mM ATP. After drying by aspirationand blotting, the plate is air dried in an incubator at 37° C.overnight. Packard TopCount adapter (Millipore) is then attached to theplate followed with addition of 120 μL Microscint 20 scintillationcocktail (Perkin Elmer) in each well. After the plate sealing, theradioactivity of the product is determined by scintillation counting onTopcount (Perkin-Elmer). IC₅₀ determination is performed by fitting thecurve of percent control activity versus the log of the inhibitorconcentration using the GraphPad Prism 3.0 software.

Example A3: PI3Kδ Scintillation Proximity Assay Materials

[γ-³³P]ATP (10 mCi/mL) was purchased from Perkin-Elmer (Waltham, Mass.).Lipid kinase substrate, D-myo-Phosphatidylinositol 4,5-bisphosphate(PtdIns(4,5)P2)D (+)-sn-1,2-di-O-octanoylglyceryl, 3-O-phospho linked(PIP2), CAS 204858-53-7, was purchased from Echelon Biosciences (SaltLake City, Utah). PI3Kδ (p110δ/p85α) was purchased from Millipore(Bedford, Mass.). ATP, MgCl₂, DTT, EDTA, MOPS and CHAPS were purchasedfrom Sigma-Aldrich (St. Louis, Mo.). Wheat Germ Agglutinin (WGA) YSi SPAScintillation Beads was purchased from GE healthcare life sciences(Piscataway, N.J.).

The kinase reaction was conducted in polystyrene 384-well matrix whiteplate from Thermo Fisher Scientific in a final volume of 25 μL.Inhibitors were first diluted serially in DMSO and added to the platewells before the addition of other reaction components. The finalconcentration of DMSO in the assay was 0.5%. The PI3K assays werecarried out at room temperature in 20 mM MOPS, pH 6.7, 10 mM MgCl₂, 5 mMDTT and CHAPS 0.03%. Reactions were initiated by the addition of ATP,the final reaction mixture consisted of 20 μM PIP2, 20 μM ATP, 0.2 μCi[γ-³³P] ATP, 4 nM PI3Kδ. Reactions were incubated for 210 min andterminated by the addition of 40 μL SPA beads suspended in quenchbuffer: 150 mM potassium phosphate pH 8.0, 20% glycerol. 25 mM EDTA, 400μM ATP. The final concentration of SPA beads was 1.0 mg/mL. After theplate sealing, plates were shaken overnight at room temperature andcentrifuged at 1800 rpm for 10 minutes, the radioactivity of the productwas determined by scintillation counting on Topcount (Perkin-Elmer).IC₅₀ determination was performed by fitting the curve of percent controlactivity versus the log of the inhibitor concentration using theGraphPad Prism 3.0 software. IC₅₀ data for the Examples is presented inTable 2 as determined by Assay A3. IC₅₀ data for Examples 361 and 363 isshown in Table 3 as determined by Assay A2.

TABLE 2 Example # PI3Kδ SPA IC₅₀ (nM)*  1 +  2 +  3 +  4 +  5 +  6 + 7 +  8 +  9 +  10 +  11 +  13 +  14 +  16 +  17 +  18 +  19 +  20(1^(st) peak) +  20 (2^(nd) peak) +++  21 +  22 +  23 +  24 +  40 ++  41+++  43 +  44 +  65 +  66 +  67 (1^(st) peak) +  68 (1^(st) peak) + 71 +  72 +  94 +  95 +  96 +  99 + 102 + 104 + 105 + 108 + 110 + 113 +115 + 118 + 121 + 139 (1^(st) peak) + 140 + 141 + 149 + 156 + 158 +159 + 161 + 163 + 164 + 166 + 167 + 168 + 169 + 183 + 184 + 188 + 189 +192 ++ 195 + 200 + 203 + 208 + 209 ++ 212 + 213 + 219 + 220 + 236 +237 + 239 + 247 + 261 + 262 + 268 + 269 + 272 + 273 + 281 + 285 + 289 +292 + 293 + 296 + 298 (1^(st) peak) + 307 + 315 + 316 + 318 + 319 +320 + 321 (1st peak) + 322 (1^(st) peak) + 310 + 311 + 323 (1^(st)peak) + 323 (2^(nd) peak) + 323 (3^(rd) peak) +++ 323 (4^(th) peak) +324 (1^(st) peak) +++ 324 (2^(nd) peak) + 325 + 345 +++ 346 + 347 + 348+++ 349 + 350 +++++ 351 +++ 352 + 353 +++++ 354 + 355 +++ 356 +++ 357 +358 +++++ 362 +

TABLE 3 Example # PI3Kδ IC₅₀ (nM)* 361 +++++ 363 +++* column symbols (for Tables 2 and 3):+ refers to <10 nM++ refers to >10 nM to 50 nM+++ refers to >50 nM to 200 nM++++ refers to >200 nM to 500 nM+++++ refers to >500 nM

Example B1: B Cell Proliferation Assay

To acquire B cells, human PBMC are isolated from the peripheral blood ofnormal, drug free donors by standard density gradient centrifugation onFicoll-Hypague (GE Healthcare, Piscataway, N.J.) and incubated withanti-CD19 microbeads (Miltenyi Biotech, Auburn, Calif.). The B cells arethen purified by positive immunosorting using an autoMacs (MiltenyiBiotech) according to the manufacture's instruction.

The purified B cells (2×10⁵/well/200 μL) are cultured in 96-wellultra-low binding plates (Corning, Corning, N.Y.) in RPMI1640, 10% FBSand goat F(ab′)2 anti-human IgM (10 μg/ml) (Invitrogen, Carlsbad,Calif.) in the presence of different amount of test compounds for threedays. [³H]-thymidine (1 μCi/well) (PerkinElmer, Boston, Mass.) in PBS isthen added to the B cell cultures for an additional 12 hours before theincorporated radioactivity is separated by filtration with water throughGF/B filters (Packard Bioscience, Meriden, Conn.) and measured by liquidscintillation counting with a TopCount (Packard Bioscience).

Example B2: Pfeiffer Cell Proliferation Assay

Pfeiffer cell line (diffuse large B cell lymphoma) are purchased fromATCC (Manassas, Va.) and maintained in the culture medium recommended(RPMI and 10% FBS). To measure the anti-proliferation activity of thecompounds, the Pfeiffer cells are plated with the culture medium (2×10³cells/well/per 200 μl) into 96-well ultra-low binding plates (Corning,Corning, N.Y.), in the presence or absence of a concentration range oftest compounds. After 3-4 days, [³H]-thymidine (1 μCi/well)(PerkinElmer, Boston, Mass.) in PBS is then added to the cell culturefor an additional 12 hours before the incorporated radioactivity isseparated by filtration with water through GF/B filters (PackardBioscience, Meridenj, Conn.) and measured by liquid scintillationcounting with a TopCount (Packard Bioscience). IC₅₀ data for selectcompounds is presented in Table 4.

TABLE 4 Example # Pfeiffer IC₅₀ (nM)*  67 (1^(st) peak) +  68 (1^(st)peak) +  96 + 102 + 104 ++ 121 ++ 139 (1^(st) peak) + 140 + 149 + 163 ++167 + 195 + 200 + 213 + 219 + 220 + 262 + 268 + 269 + 315 + 354 + 357 +346 + 347 + 349 + *column symbols: + refers to ≤10 nM ++ refers to >10nM to 50 nM

Example C: Akt Phosphorylation Assay

Ramos cells (B lymphocyte from Burkitts lymphoma) are obtained from ATCC(Manassas, Va.) and maintained in RPMI1640 and 10% FBS. The cells (3×10⁷cells/tube/3 mL in RPMI) are incubated with different amounts of testcompounds for 2 hrs at 37° C. and then stimulated with goat F(ab′)2anti-human IgM (5 μg/mL) (Invitrogen) for 17 minutes in a 37° C. waterbath. The stimulated cells are spun down at 4° C. with centrifugationand whole cell extracts are prepared using 300 μL lysis buffer (CellSignaling Technology, Danvers, Mass.). The resulting lysates aresonicated and supernatants are collected. The phosphorylation level ofAkt in the supernatants are analyzed by using PathScan phospho-Akt1(Ser473) sandwich ELISA kits (Cell Signaling Technology) according tothe manufacturer's instruction.

Example D: Pfeiffer Model of Lymphoma

Methods:

Female SCID mice, (5 to 8 weeks of age, Charles River Laboratories,Wilmington, Mass.) were inoculated with 1×107 tumor cells (Pfeiffer,ATCC #CRL-2632, Manassas, Va.) and matrigel (BD Biosciences #354234) in0.2 mL sterile saline. The inoculation was performed subcutaneously onthe flank. Tumor tissue fragments (approximately 3 mm×3 mm) werecollected 3 to 6 weeks after the inoculation of cultured cells andimplanted subcutaneously in lieu of cellular inoculation. Tissuefragments were implanted as solid pieces using blunt-tip forceps. Thetreatment of tumor bearing mice was started 15 to 25 days after tumorinoculation, depending upon the tumor size. Animals were sorted toobtain roughly equivalent mean tumor volumes in each group. Minimum meantumor volume in all groups was 150 mm3 on the first day of treatment andgroups consisted of 7 animals. Experimental therapeutic agent, Example347, was administered to mice orally (PO). Treatment frequency was 2times daily for a minimum of 14 days for efficacy. The size ofsubcutaneous tumors was measured 2 to 3 times weekly using a digitalcaliper. The tumor volume was calculated by measuring the tumor in 2dimensions and utilizing the equation: Volume=[Length×(Width2)]/2; wherethe larger number was length, and the smaller number width. If multipletumors were formed, the final volume was the sum of the individualtumors subject to the same equation: eg, 2 tumors;Volume={[L1×(W1)2]/2}+{[L2×(W2)2]/2}. Effects on tumor growth werereported as percent tumor growth inhibition (% TGI). Percent TGI wascalculated with the equation: (1−(Tx vol./control vol.))*100, wherecontrol volume was the vehicle or untreated tumor volume on a given day,and Tx volume was any treatment group tumor volume on that same day.Statistical differences between treatment and vehicle controls wereassessed using ANOVA: Single Factor test.

Results:

Example 347 was evaluated as a single agent in the Pfeiffer human tumorxenograft model of diffuse large B-cell lymphoma, a subtype of NHL.Pfeiffer cancer cells were shown to be sensitive to theanti-proliferative effects of Example 347 in vitro. Therefore, a tumormodel was established based on subcutaneous inoculation of tumor cellsinto immune compromised SCID mice and tumor-bearing mice received twicedaily oral doses of vehicle or Example 347 at 0.3, 1, 3, or 10 mg/kg for14 days. Example 347 treatment inhibited tumor growth by 22%, 24%, 36%,and 58% (percent tumor growth inhibition) with increasing dose (FIG. 2).

Various modifications of the invention, in addition to those describedherein, will be apparent to those skilled in the art from the foregoingdescription. Such modifications are also intended to fall within thescope of the appended claims. Each reference, including all patent,patent applications, and publications, cited in the present applicationis incorporated herein by reference in its entirety.

1. A method of treating a disease selected from autoimmune hemolyticanemia pemphigus, and myelofibrosis in a patient, comprisingadministering to said patient a therapeutically effective amount of acompound of Formula I:

or a pharmaceutically acceptable salt thereof, wherein: R² is C₁₋₆ alkylor C₁₋₆ haloalkyl; R⁴ is halo, OH, CN, C₁₋₄ alkyl, C₁₋₄ haloalkyl, C₁₋₄alkoxy, or C₁₋₄ haloalkoxy; R⁵ is halo, OH, CN, C₁₋₄ alkyl, C₁₋₄haloalkyl, C₁₋₄ alkoxy, or C₁₋₄ haloalkoxy; Cy is selected from C₃₋₇cycloalkyl, 4-7 membered heterocycloalkyl, phenyl, and 5-6 memberedheteroaryl, each of which is optionally substituted with 1, 2, 3, or 4independently selected R³ groups; each R³ is independently selected fromCy¹, —(C₁₋₃ alkylene)-Cy¹, halo, CN, NO₂, C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆alkynyl, C₁₋₆ haloalkyl, OR^(a1), SR^(a1), C(═O)R^(b1),C(═O)NR^(c1)R^(d1), C(═O)OR^(a1), OC(═O)R^(b1)OC(═O)NR^(c1)R^(d1),NR^(c1)R^(d1), NR^(c1)C(═O)R^(b1), NR^(c1)C(═O)OR^(b1),NR^(c1)C(═O)NR^(c1)R^(d1), C(═NR^(e))R^(b1), C(═NR^(e))NR^(c1)R^(d1),NR^(c1)C(═NR^(e))NR^(c1)R^(d1), NR^(c1)S(═O)R^(b1),NR^(c1)S(═O)₂NR^(c1)R^(d1), S(═O)R^(b1), S(═O)₂R^(b1), andS(═O)₂NR^(c1)R^(d1); wherein said C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆ alkynylare each optionally substituted with 1, 2, 3, or 4 independentlyselected R¹¹ groups; each Cy¹ is independently selected from C₃₋₇cycloalkyl, 4-7 membered heterocycloalkyl, phenyl, and 5-6 memberedheteroaryl, each of which is optionally substituted with 1, 2, 3, or 4independently selected R¹¹ groups; each R^(a1), R^(c1), and R^(d1) isindependently selected from H, C₁₋₆ alkyl, C₁₋₆ haloalkyl, C₂₋₆ alkenyl,C₂₋₆ alkynyl, C₃₋₇ cycloalkyl, 4-7 membered heterocycloalkyl, phenyl,and 5-6 membered heteroaryl; wherein said C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆alkynyl, C₃₋₇ cycloalkyl, 4-7 membered heterocycloalkyl, phenyl and 5-6membered heteroaryl are each optionally substituted with 1, 2, or 3independently selected R¹¹ groups; each R^(b1) is independently selectedfrom C₁₋₆ alkyl, C₁₋₆ haloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇cycloalkyl, 4-7 membered heterocycloalkyl, phenyl, and 5-6 memberedheteroaryl; wherein said C₁₋₆ alkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, C₃₋₇cycloalkyl, 4-7 membered heterocycloalkyl, phenyl, and 5-6 memberedheteroaryl are each optionally substituted with 1, 2, or 3 independentlyselected R¹¹ groups; or R^(c1) and R^(d1) together with the N atom towhich they are attached form a 4-, 5-, 6-, or 7 memberedheterocycloalkyl group, which is optionally substituted with —OH or C₁₋₃alkyl; each R^(e) is independently selected from H, CN, OH, C₁₋₄ alkyl,and C₁₋₄ alkoxy; and each R¹¹ is independently selected from OH, NO₂,CN, halo, C₁₋₃ alkyl, C₂₋₃ alkenyl, C₂₋₃ alkynyl, C₁₋₃ haloalkyl,cyano-C₁₋₃ alkyl, HO—C₁₋₃ alkyl, C₁₋₃ alkoxy-C₁₋₃ alkyl, C₃₋₇cycloalkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkoxy, amino, C₁₋₃ alkylamino,di(C₁₋₃ alkyl)amino, thio, C₁₋₃ alkylthio, C₁₋₃ alkylsulfinyl, C₁₋₃alkylsulfonyl, carbamyl, C₁₋₃ alkylcarbamyl, di(C₁₋₃ alkyl)carbamyl,carboxy, C₁₋₃ alkylcarbonyl, C₁₋₄ alkoxycarbonyl, C₁₋₃alkylcarbonylamino, C₁₋₃ alkylsulfonylamino, aminosulfonyl, C₁₋₃alkylaminosulfonyl, di(C₁₋₃ alkyl)aminosulfonyl, aminosulfonylamino,C₁₋₃ alkylaminosulfonylamino, di(C₁₋₃ alkyl)aminosulfonylamino,aminocarbonylamino, C₁₋₃ alkylaminocarbonylamino, and di(C₁₋₃alkyl)aminocarbonylamino.
 2. The method of claim 1, wherein R² is C₁₋₃alkyl or C₁₋₃ fluoroalkyl.
 3. The method of claim 1, wherein R² ismethyl, ethyl, or 2,2-difluoromethyl.
 4. The method of claim 1, whereinR² is methyl.
 5. The method of claim 1, wherein R² is ethyl.
 6. Themethod of claim 1, wherein R⁴ is halo, CN, or C₁₋₃ alkyl.
 7. The methodof claim 1, wherein R⁴ is F, Cl, CN, or methyl.
 8. The method of claim1, wherein R⁴ is F.
 9. The method of claim 1, wherein R⁴ is Cl.
 10. Themethod of claim 1, wherein R⁴ is CN.
 11. The method of claim 1, whereinR⁴ is methyl.
 12. The method of claim 1, wherein R⁵ is halo, CN, or C₁₋₃alkyl.
 13. The method of claim 1, wherein R⁵ is Cl, CN, or methyl. 14.The method of claim 1, wherein R⁵ is Cl.
 15. The method of claim 1,wherein R⁵ is CN.
 16. The method of claim 1, wherein R⁵ is methyl. 17.The method of claim 1, wherein Cy is selected from C₃₋₆ cycloalkyl, 4-6membered heterocycloalkyl, phenyl, and 5-6 membered heteroaryl, each ofwhich is optionally substituted with 1, 2, 3, or 4 independentlyselected R³ groups.
 18. The method of claim 1, wherein Cy is 4-6membered heterocycloalkyl, each of which is optionally substituted with1, 2, 3, or 4 independently selected R³ groups.
 19. The method of claim1, wherein Cy is selected from a cyclopropyl ring, a phenyl ring, anazetidine ring, a pyrrolidine ring, a piperidine ring,3-oxo-morpholin-6-yl, 2-oxo-pyrrolidin-4-yl, 2-oxo-oxazolidin-4-yl,2-oxo-oxazolidin-5-yl, a pyrazole ring, a pyridine ring, and apyrimidine ring, each of which is optionally substituted with 1, 2, 3,or 4 independently selected R³ groups.
 20. The method of claim 1,wherein: each R³ is independently selected from Cy¹, —(C₁₋₃alkylene)-Cy¹, halo, CN, C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1),C(═O)R^(b1), C(═O)OR^(a1), C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), whereinsaid C₁₋₆ alkyl is optionally substituted with 1, 2, 3, or 4independently selected R¹¹ groups; each Cy¹ is independently C₃₋₇cycloalkyl, which is optionally substituted with 1, 2, 3, or 4independently selected R¹¹ groups; each R^(a1), R^(c1), and R^(d1) isindependently selected from H and C₁₋₆ alkyl, wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, or 3 independently selected R¹¹groups; each R^(b1) is independently C₁₋₆ alkyl, which is optionallysubstituted with 1, 2, or 3 independently selected R¹¹ groups; and eachR¹¹ is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃ haloalkoxy,amino, C₁₋₃ alkylamino, di(C₁₋₃ alkyl)amino, C₁₋₃ alkylcarbonyl, C₁₋₃alkoxycarbonyl, carbamyl, C₁₋₃ alkylcarbamyl, or di(C₁₋₃ alkyl)carbamyl.21. The method of claim 1, wherein: R² is C₁₋₃ alkyl or C₁₋₃fluoroalkyl; R⁴ is halo, CN, or C₁₋₃ alkyl; R⁵ is halo, CN, or C₁₋₃alkyl; Cy is selected from C₃₋₆ cycloalkyl, 4-6 memberedheterocycloalkyl, phenyl, and 5-6 membered heteroaryl, each of which isoptionally substituted with 1, 2, 3, or 4 independently selected R³group; each R³ is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹,halo, CN, C₁₋₆ alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups; each Cy¹ is independently C₃₋₇ cycloalkyl, which is optionallysubstituted with 1, 2, 3, or 4 independently selected R¹¹ groups; eachR^(a1), R^(c1), and R^(d1) is independently selected from H and C₁₋₆alkyl, wherein said C₁₋₆ alkyl is optionally substituted with 1, 2, or 3independently selected R¹¹ groups; each R^(b1) is independently C₁₋₆alkyl, which is optionally substituted with 1, 2, or 3 independentlyselected R¹¹ groups; and each R¹¹ is independently OH, CN, halo,cyano-C₁₋₃ alkyl, C₁₋₃ haloalkoxy, amino, C₁₋₃ alkylamino, di(C1-3alkyl)amino, C₁₋₃ alkylcarbonyl, C₁₋₃ alkoxycarbonyl, carbamyl, C₁₋₃alkylcarbamyl, or di(C₁₋₃ alkyl)carbamyl.
 22. The method of claim 1,wherein: R² is methyl, ethyl, or 2,2-difluoromethyl; R⁴ is F, Cl, CN, ormethyl; R⁵ is Cl, CN, or methyl; Cy is selected from C₃₋₆ cycloalkyl,4-6 membered heterocycloalkyl, phenyl, and 5-6 membered heteroaryl, eachof which is optionally substituted with 1, 2, 3, or 4 independentlyselected R³ group; each R³ is independently selected from Cy¹, —(C₁₋₃alkylene)-Cy¹, halo, CN, C₁₋₆ alkyl, OR^(a), NR^(c1)R^(d1), C(═O)R^(b1),C(═O)OR^(a1), C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆alkyl is optionally substituted with 1, 2, 3, or 4 independentlyselected R¹¹ groups; each Cy¹ is independently C₃₋₇ cycloalkyl, which isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups; each R^(a1), R^(c1), and R^(d1) is independently selected from Hand C₁₋₆ alkyl, wherein said C₁₋₆ alkyl is optionally substituted with1, 2, or 3 independently selected R¹¹ groups; each R^(b1) isindependently C₁₋₆ alkyl, which is optionally substituted with 1, 2, or3 independently selected R¹¹ groups; and each R¹¹ is independently OH,CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃ haloalkoxy, amino, C₁₋₃ alkylamino,di(C₁₋₃ alkyl)amino, C₁₋₃ alkylcarbonyl, C₁₋₃ alkoxycarbonyl, carbamyl,C₁₋₃ alkylcarbamyl, or di(C₁₋₃ alkyl)carbamyl.
 23. The method of claim1, wherein: R² is methyl, ethyl, or 2,2-difluoromethyl; R⁴ is F, Cl, CN,or methyl; R⁵ is Cl, CN, or methyl; Cy is selected from a cyclopropylring, a phenyl ring, an azetidine ring, a pyrrolidine ring, a piperidinering, 3-oxo-morpholin-6-yl, 2-oxo-pyrrolidin-4-yl,2-oxo-oxazolidin-4-yl, 2-oxo-oxazolidin-5-yl, a pyrazole ring, apyridine ring, and a pyrimidine ring, each of which is optionallysubstituted with 1, 2, 3, or 4 independently selected R³ groups each R³is independently selected from Cy¹, —(C₁₋₃ alkylene)-Cy¹, halo, C₁₋₆alkyl, OR^(a1), NR^(c1)R^(d1), C(═O)R^(b1), C(═O)OR^(a1),C(═O)NR^(c1)R^(d1), and S(═O)₂R^(b1), wherein said C₁₋₆ alkyl isoptionally substituted with 1, 2, 3, or 4 independently selected R¹¹groups; each Cy¹ is independently selected from cyclopropyl andcyclobutyl, each of which is optionally substituted with 1, 2, 3, or 4independently selected R¹¹ groups; each R^(a1), R^(c1), and R^(d1) isindependently selected from H and C₁₋₄ alkyl; wherein said C₁₋₄ alkyl isoptionally substituted with 1, 2, or 3 independently selected R¹¹groups; each R^(b1) is independently C₁₋₄ alkyl, which is optionallysubstituted with 1, 2, or 3 independently selected R¹¹ groups; each R¹¹is independently OH, CN, halo, cyano-C₁₋₃ alkyl, C₁₋₃ alkoxy-C₁₋₃ alkyl,C₁₋₃ alkoxy, C₁₋₃ haloalkoxy, amino, C₁₋₃ alkylamino, di(C₁₋₃alkyl)amino, C₁₋₃ alkylcarbonyl, C₁₋₄ alkoxycarbonyl, carbamyl, C₁₋₃alkylcarbamyl, or di(C₁₋₃ alkyl)carbamyl.
 24. The method of claim 1,wherein the compound is a compound of Formula II:

or a pharmaceutically acceptable salt thereof.
 25. The method of claim1, wherein the compound is a compound of Formula III:

or a pharmaceutically acceptable salt thereof.
 26. The method of claim1, wherein the compound is a compound of Formula IV:

or a pharmaceutically acceptable salt thereof, wherein: G is NH, n is 1,and V is O; or G is NH, n is 0, and V is O or CH₂; or G is O, n is 0 andV is NH.
 27. The method of claim 1, wherein the compound is a compoundof Formula IVa:

or a pharmaceutically acceptable salt thereof.
 28. The method of claim1, wherein the compound is a compound of Formula IVb:

or a pharmaceutically acceptable salt thereof.
 29. The method of claim1, wherein the compound is a compound of Formula IVc:

or a pharmaceutically acceptable salt thereof.
 30. The method of claim1, wherein the compound is a compound of Formula IVd:

or a pharmaceutically acceptable salt thereof.
 31. (canceled)
 32. Themethod of claim 1, or a pharmaceutically acceptable salt thereof,wherein the starred carbon in Formula I:

is a chiral carbon and said compound or said salt is the (S)-enantiomer.33. The method of claim 1, wherein the compound is selected from:1-{1-[5-Chloro-3-(1-isopropylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[3-(1-Acetylazetidin-3-yl)-5-chloro-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[5-Chloro-2-methoxy-4-methyl-3-(1-propionylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-(1-{5-Chloro-3-[1-(cyclopropylmethyl)azetidin-3-yl]-2-methoxy-4-methylphenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[5-chloro-2-methoxy-4-methyl-3-(1-methylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[5-Chloro-3-(1-ethylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[5-Chloro-3-(1-isobutylazetidin-3-yl)-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[3-(1-sec-butylazetidin-3-yl)-5-chloro-2-methoxy-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-(1-{5-Chloro-2-methoxy-3-[1-(2-methoxyethyl)azetidin-3-yl]-4-methylphenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-methylazetidine-1-carboxamide;5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide;1-{1-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide;4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide;4-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N-methylpicolinamide;4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)pyridine-2-carboxamide;4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide;2-(4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-1H-pyrazol-1-yl)ethanol;3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-N,N,6′-trimethylbiphenyl-4-carboxamide;3′-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5′-chloro-3-fluoro-2′-methoxy-N,6′-dimethylbiphenyl-4-carboxamide;5-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N-(2-hydroxyethyl)picolinamide;4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide;4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-cyano-2-methoxy-6-methylphenyl}-N-(2-hydroxyethyl)-N-methylpyridine-2-carboxamide;5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[4,3-c]pyridin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[5-(methylsulfonyl)pyridin-3-yl]benzonitrile;5-(3-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-6-cyano-2-ethoxyphenyl)-N,N-dimethylpicolinamide5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylnicotinamide5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-N,N-dimethylpyridine-2-carboxamide;1-{1-[5-Chloro-4-fluoro-3-(1-isopropylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidin-1-yl)propan-2-ol;2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-fluoro-2-methoxyphenyl}azetidin-1-yl)ethanol;1-{1-[5-Chloro-4-fluoro-2-methoxy-3-(1-oxetan-3-ylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-{1-[5-Chloro-2-ethoxy-3-(1-isopropylazetidin-3-yl)-4-methylphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)ethanol;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)propan-2-ol;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-ol;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propan-2-ol;2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)ethanol;(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)acetonitrile;1-(1-{5-Chloro-2-methoxy-4-methyl-3-[1-(2,2,2-trifluoroethyl)azetidin-3-yl]phenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-N-methylpropanamide;2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-3,3,3-trifluoropropan-1-ol;(2R)-3-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1,1,1-trifluoropropan-2-ol;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-ol;(2R)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-1-oxopropan-2-ol;1-{1-[5-Chloro-2-ethoxy-4-fluoro-3-(1-isopropylazetidin-3-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidin-1-yl)-2-methylpropan-2-ol;1-(1-{5-Chloro-2-ethoxy-4-fluoro-3-[1-(2,2,2-trifluoroethyl)azetidin-3-yl]phenyl}ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;(2S)-1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}azetidin-1-yl)propan-2-ol;(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)propan-1-ol;1-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-2-methylpropan-2-ol;(2R)-2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)-N,N-dimethylpropanamide;[1-{3-3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}azetidin-1-yl)cyclobutyl]acetonitrile;1-{1-[5-Chloro-2-methoxy-4-methyl-3-(1-methylpiperidin-4-yl)phenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-(4-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}piperidin-1-yl)-2-methylpropan-2-ol;3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}cyclobutanol;5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamide;1-[1-(5-Chloro-3-{1-[2-(dimethylamino)ethyl]-1H-pyrazol-4-yl}-2-methoxy-4-methylphenyl)ethyl]-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;2-[(5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}pyridin-2-yl)amino]ethanol;2-(5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)pyridin-2-yloxy)ethanol;5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-(2,2-difluoroethoxy)-6-methylphenyl)-N,N-dimethylpicolinamide;5-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-methylphenyl)-N,N-dimethylpicolinamide;4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-methoxy-6-methylphenyl)-N,N-dimethylpicolinamide;2-(4-(3-(1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-methylphenyl)-1H-pyrazol-1-yl)acetamide;6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}-N,N-dimethylnicotinamide;5-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-4-methoxy-2-methyl-3-(1-methyl-1H-pyrazol-4-yl)benzonitrile;1-(1-(5-chloro-2-methoxy-4-methyl-3-(pyrimidin-5-yl)phenyl)ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;1-(1-(3-(2-aminopyrimidin-5-yl)-5-chloro-2-methoxy-4-methylphenyl)ethyl)-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-ethoxybenzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(1-methylazetidin-3-yl)benzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxyethyl)azetidin-3-yl]benzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}benzonitrile;tert-Butyl2-(3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanoate;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxy-1,1-dimethylethyl)azetidin-3-yl]benzonitrile;2-(3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-ethoxyphenyl}azetidin-1-yl)-2-methylpropanamide;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[1-(2-hydroxy-2-methylpropanoyl)azetidin-3-yl]benzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-azetidin-3-yl-6-chloro-3-methoxybenzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxy-2-(1-methylazetidin-3-yl)benzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-[1-(2-hydroxyethyl)azetidin-3-yl]-3-methoxybenzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}-3-methoxybenzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-2-[1-(2-hydroxy-1-methylethyl)azetidin-3-yl]-3-methoxybenzonitrile;2-(1-Acetylazetidin-3-yl)-4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxybenzonitrile;4-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-methoxy-2-[1-(methylsulfonyl)azetidin-3-yl]benzonitrile;Methyl3-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxylate;3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}-N-(tert-butyl)azetidine-1-carboxamide;3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}azetidine-1-carboxamide;3-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-6-cyano-2-methoxyphenyl}-N,N-dimethylazetidine-1-carboxamide;1-{1-[4,5-Dichloro-3-(1-ethylazetidin-3-yl)-2-methoxyphenyl]ethyl}-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-4-amine;4-[1-(4-Amino-5-methyl-7H-pyrrolo[2,3-d]pyrimidin-7-yl)ethyl]-6-chloro-3-ethoxy-2-(1-isopropylazetidin-3-yl)benzonitrile;4-[-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-2-{1-[(2S)-2-hydroxypropyl]azetidin-3-yl}-3-methoxy-6-methylbenzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-[6-(1-hydroxy-1-methylethyl)pyridin-3-yl]benzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-pyrrolidin-1-ylbenzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(3-methoxyazetidin-1-yl)benzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-(1-isopropylazetidin-3-yl)-6-methylbenzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-[1-(2-hydroxy-2-methylpropyl)azetidin-3-yl]-6-methylbenzonitrile;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-3-ethoxy-2-[1-(2-hydroxy-2-methylpropanoyl)azetidin-3-yl]-6-methylbenzonitrile;4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-methylphenyl}pyrrolidin-2-one;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(2-oxo-1,3-oxazolidin-5-yl)benzonitrile;6-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}morpholin-3-one;5-{3-[1-(4-Amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-methoxy-6-methylphenyl}-1,3-oxazolidin-2-one;4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}pyrrolidin-2-one;4-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-6-chloro-3-ethoxy-2-(5-oxopyrrolidin-3-yl)benzonitrile;4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one;5-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}-1,3-oxazolidin-2-one;and4-(1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-6-chloro-2-(1-(2-hydroxypropyl)azetidin-3-yl)-3-methoxybenzonitrile;or a pharmaceutically acceptable salt of any of the aforementioned.34-36. (canceled)
 37. The method of claim 1, wherein the compound is4-{3-[1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl]-5-chloro-2-ethoxy-6-fluorophenyl}pyrrolidin-2-one,or a pharmaceutically acceptable salt thereof.
 38. The method of claim1, wherein the compound is(S)-4-(3-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one,or a pharmaceutically acceptable salt thereof.
 39. The method of claim1, wherein the compound is(R)-4-(3-((S)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one,or a pharmaceutically acceptable salt thereof.
 40. The method of claim1, wherein the compound is(S)-4-(3-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one,or a pharmaceutically acceptable salt thereof.
 41. The method of claim1, wherein the compound is(R)-4-(3-((R)-1-(4-amino-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)ethyl)-5-chloro-2-ethoxy-6-fluorophenyl)pyrrolidin-2-one,or a pharmaceutically acceptable salt thereof.
 42. The method of claim1, wherein the disease is autoimmune hemolytic anemia.
 43. The method ofclaim 37, wherein the disease is autoimmune hemolytic anemia.
 44. Themethod of claim 38, wherein the disease is autoimmune hemolytic anemia.45. The method of claim 39, wherein the disease is autoimmune hemolyticanemia.
 46. The method of claim 40, wherein the disease is autoimmunehemolytic anemia.
 47. The method of claim 41, wherein the disease isautoimmune hemolytic anemia.
 48. The method of claim 1, wherein thedisease is pemphigus.
 49. The method of claim 37, wherein the disease ispemphigus.
 50. The method of claim 38, wherein the disease is pemphigus.51. The method of claim 39, wherein the disease is pemphigus.
 52. Themethod of claim 40, wherein the disease is pemphigus.
 53. The method ofclaim 41, wherein the disease is pemphigus.
 54. The method of claim 1,wherein the disease is myelofibrosis.
 55. The method of claim 37,wherein the disease is myelofibrosis.
 56. The method of claim 38,wherein the disease is myelofibrosis.
 57. The method of claim 39,wherein the disease is myelofibrosis.
 58. The method of claim 40,wherein the disease is myelofibrosis.
 59. The method of claim 41,wherein the disease is myelofibrosis.